Doing scientific research in post-Soviet Russia is challenging but there are solutions that could prevent the massive brain drain witnessed in recent decades A scientist living in Russia is often asked two questions: “Why haven't you left?” and “Is it still possible to work there?” The best response to the first question is, “Why should I?”—which either terminates the conversation or leads to a stimulating discussion about the fate of the world. The second question, however, deserves a serious answer. In fact, this is the question that every one of us keeps asking ourselves.There is no simple answer. The biggest problems we face are brain drain, inadequate infrastructure, and lack of money . In the Soviet Union, fundamental science was supported to a great extent by military expenditure. Thus, it is not surprising that Soviet physics and mathematics were more successful than other fields, such as biology. In the 1990s, military spending on science declined sharply, although the exact numbers are hard to estimate. This year, the direct funding of science constitutes only 1.78% of Russia's national budget , although the law stipulates that this figure should be at least 4%. Still, this funding amounts to 46.2 billion rubles (approximately US$1.6 billion), more than twice the amount spent in 2000. Although this figure looks negligible compared with spending on science in the United States and many European countries, it could still be sufficient to support existing actively working groups at a reasonable level.There are several mechanisms for distributing funds for research. The major share comes via Russia's Department of Science and the Russian Academy of Sciences. The Academy, unlike its Western analogs, not only acts as a consulting body of experts, but also has the authority to distribute money . The fMoney is also distributed through the Russian Foundation for Basic Research (RFBR). The decisionmaking mechanism used by RFBR is closer to Western standards, and involves anonymous refereeing followed by board discussions. Although its grants are rather small , they provide important additional support for many small and mediumsized groups that may receive several such grants for different projects. In addition, RFBR supports the publication and translation of books, travel to international conferences, the organization of conferences in Russia, and similar activities. Unfortunately, several programs  have recently been transferred from RFBR to a newly established government office, and have thus become less independent.International collaboration and research grants are a major source of support for many active research groups . SeveralAnother source of financial support is direct collaboration between Western and Russian laboratories. Even after a relatively short visit, the salary of a visiting researcher abroad can be stretched for several months back home in Russia; even more importantly, experimental biologists visiting foreign labs have access to modern instruments and chemicals, which allows them to do modern research. The hosts of such visits are often  recent immigrants from Russia, and in such cases the collaborations may have roots in older days .As well as supporting Russian science directly, international collaboration plays an important indirect role because it is less influenced by local politics. In fact, one of the main positive impacts of the New York–based International Science Foundation set up by George Soros in early 1990s was that it demonstrated the possibility of open competition with clearly defined rules—something unheard of in Soviet times—and thus served as a model for the RFBR, which was organized at approximately the same time.Unfortunately, international ties, especially with the United States, have been adversely affected by recent changes in visa procedures, which have become lengthy (leading to many missed conferences) and, even worse, completely unpredictable e.g., . The grahttp://www.evrogen.com/about.shtml).The demand for fundamental and even for applied biological research from Russian industry is almost nonexistent. The pharmaceutical industry is content to produce generics, while Russian biotech companies are still exploiting old strains developed in the Soviet Union. However, some laboratories are conducting outsourced research, and there are now research outposts of Western and Japanese companies in Russia organized as standard industrial labs. On one hand, this work is a dead end for Russian scientists, because the results of such research normally cannot be published. This is a serious problem, especially for young scientists who want to establish themselves. On the other hand, royalties from patents or commercialization of the products can be used to support further research. One group that has followed this path successfully is Sergey Lukyanov's lab at the Shemyakin Institute of Bioorganic Chemistry in Moscow, Russia. They have developed the subtractive hybridization technique for enrichment of clone libraries by rare transcripts or specific genomic fragments , and areAnother major problem is the degradation of infrastructure. Only a few labs can afford modern equipment and instruments, and for many others, even standard chemicals are too expensive. This leads to a vicious circle: without equipment, a lab cannot conduct experiments at the level demanded by highimpact journals—and without such publications, it cannot compete for large grants. Smaller RFBR grants, while simpler to obtain, are insufficient to purchase large pieces of equipment, and funds from several grants or several years cannot be combined due to bureaucratic restrictions. Thus, the only hope for these labs, apart from international collaboration, is a personal connection with senior bureaucrats that might result in an (un)expected windfall.Having the funds to purchase modern equipment abroad is only the first hurdle; the many conflicting rules and restrictions, inefficiency, and corruption within the system can subsequently hold up the process. Some items, such as tissue samples or animals, are virtually impossible to import legally. The process of clearing the shipments through customs is a difficult, timeconsuming job. Grigory Kopelevich, the Howard Hughes Medical Institute's Russian representative, recalls a story of a grantee whose microscope was stopped at customs because the box contained two screwdrivers not specified in the order. Fortunately, to resolve the issue, it was sufficient to present one of the screwdrivers to a customs officer as a gift.Nature, and only two to Science. More specialized journals are available only in Moscow and perhaps St. Petersburg. This is partially offset by the proliferating open-access journals from the Public Library of Science and BioMed Central, free electronic versions of older issues provided by some journals, free subscriptions for Russian academic institutes granted by some publishers or purchased by international foundations   , reprintThese problems, along with low salaries, have naturally led to a huge brain drain. Entire generations have been decimated ; the deaDespite all this, science in Russia is very much alive. Not-yet-Nobel-prizewinner Alexei Abrikosov's repeated exhorations to the scientific community “to help all the talented scientists leave Russia and to ignore the rest” were met by universal disgust . There aWhat can be done by the international community to support what is left of Russian science? Of course, direct support in the form of competitive grants is important, especially if there are few restrictions on spending; even the most carefully considered procedure cannot foresee all possible situations. But even more useful is the creation of joint research centers, such as the one opened by the international Ludwig Institute for Cancer Research (LICR) based jointly in Zurich, New York, and London, and the Belozersky Institute of Physico-Chemical Biology of Moscow State University in Moscow, Russia. This research center began with limited support for several stronger groups, and is gradually moving toward integration of a research program in Moscow with other LICR projects, and real collaboration between Moscow groups and LICR labs elsewhere.www.oncoimmunology.ru/index_e.htm), and the participation of Western scientists in scientific conferences in Russia are important not only because they provide a fresh understanding of emerging trends, but also because they create personal contacts between Russian and Western scientists that often lead to fruitful collaboration.One of the most essential elements of successful research is access to up-to-date information. Consequently, any initiative that provides open access to scientific literature and databases is extremely useful. Seminars, lecture courses , indicating that international collaboration is the most reliable source of support for top-level research. (Text and figures in this box courtesy of Alexey Kondrashov.)


Correlation between the expression levels of genes which are located close to each other on the genome has been found in various organisms, including yeast, drosophila and humans. Since such a correlation could be explained by several biochemical, evolutionary, genetic and technological factors, there is a need for statistical models that correspond to specific biological models for the correlation structure.We modelled the pairwise correlation between the expressions of the genes in a Drosophila microarray experiment as a normal mixture under Fisher's z-transform, and fitted the model to the correlations of expressions of adjacent as well as non-adjacent genes. We also analyzed simulated data for comparison. The model provided a good fit to the data. Further, correlation between the activities of two genes could, in most cases, be attributed to either of two factors: the two genes both being active in the same age group (adult or embryo), or the two genes being in proximity of each other on the chromosome. The interaction between these two factors was weak.Correlation between the activities of adjacent genes is higher than between non-adjacent genes. In the data we analyzed, this appeared, for the most part, to be a constant effect that applied to all pairs of adjacent genes. Several studies  typically spans several genes , many orphanides)), and aenes (Roy). Therefenes (Roy) suggestenes (Roy). And Chenes , or in  Llorente). Parisi Llorente found a  Llorente suggesteFinally, since the action of a transcription factor on a promotor gets weaker with distance, genes belonging to the same pathway should show stronger correlation if they are located close to each other (Dorsett ). Due toconsistent coregulation structure. Suppose that two genes  are co-regulated because, for example, they participate in the same pathway. They would, then, show a strong correlation because they would be co-regulated in all tissue samples. This need not necessarily be the case with the chromatin domain model: the segments of euchromatin in one tissue sample may overlap with those in another tissue sample. This latter scenario, we call an inconsistent coregulation structure. With consistent coregulation, adjacent gene pairs will show either strong positive correlation, or they will be uncorrelated. With inconsistent coregulation, all adjacent gene pairs will show a modest positive correlation.An important consequence of the evolution-based theories is that they predict a w be a fixed window size, e.g. 10. For each window of w adjacent genes, the average pairwise Pearson correlation coefficient within the window was computed. If that measure was found to be significant at, say, 1 - α = 0.999 , all the genes in the sequence were tagged. Doing this for all windows , the total number of tagged genes was counted. Then the experiment was repeated with shuffled genes , and the number of tagged genes in the shuffled experiment was subtracted from the number of tagged genes in the original experiment. This difference  grows with window size and starts plateauing for a window size of approximately 10. Spellman and Rubin interpreted this as evidence for gene interaction within regions of approximately that size.In a microarray analysis of gene expressions in 35 pools of drosophila embryos and 54 adult drosophilae  Correlation of gene expression that can be attributed to consistent coregulation, fromB) The uniform correlation expected under an hypothesis of inconsistent corregulation.We used the Drosophila data published by Spellman and Rubin. This daμ0), was 0.014 with a standard error of 0.002. . The fact that the standard deviation of the third component, σ+, is greater than that of the other components, suggests that the co-regulation of some gene pairs was stronger than that of others.The distribution of the correlation coefficient between gene pairs in general,  was fitted with a three-component normal mixture, based on the Arcus Tangens Hyperbolic-transform (Fisher's z) of the correlation coefficients, which was validated with a Kolmogorov-Smirnov test (p = 0.6). As shown in table μ0 and μ+ are substantially higher. This suggests that the effect of adjacency is, for the most part, a mechanism that applies to adjacent-gene pairs in general, not just to specific adjacent-gene pairs. One way to illustrate this is by means of a qq-plot , for random and adjacent gene pairs in four subsets of the flies: one containing all 54 adult pools, one containing all 35 embryo pools, and two random subsets of 35 and 54 pools, respectively. One can observe that μ0  is always high for adjacent gene pairs and low for random gene pairs. On the other hand μ+ and the "+"-fraction  are always high in heterogenous fly groups and low in homogenous one. That difference between heterogenous and homogenous groups is what we expected: In a homogenous fly group, less gene pairs will come up as co-regulated, because some pathways may either be active in all flies, or inactive in all flies. That age groups does not explain all correlation, is hardly surprising: although the adult group and the embryo group are less heterogenous than the mixed group, neither is homogenous.Table μ0 was 0.114 for the correlation between the expression of a gene and that of its direct neighbor, and 0.08 for the correlation with its second neighbor. From the fifth neighbor and until at least the tenth, a stable level of 0.03 is reached, which is still higher than for distant gene pairs. One possible interpretation of this is that two different co-regulation effects exist: a short-segment effect accounting for a correlation of approximately 0.114-0.03 = 0.081, and a long-segment effect accounting for a correlation of approximately 0.03-0.014 = 0.016. At first we had the suspicion that this stable level of 0.03 was a whole-chromosome effect, but that was not the case. We found no significant difference between the overall average correlation of random gene pairs from the same chromosome and random gene pairs from different chromosomes.To get an impression of the size of the segments involved in co-regulation, we fitted the three-component mixture to the correlation coefficient between the expressions of more distant gene pairs. μ0) for the second neighbor would be the square of that for the first neighbor. However, the unexpectedly small difference between the two values could be due to the fact that some second neighbors are physically quite close. To confirm or reject the hypothesis of a first-order process, however, the analysis must be based on physical distance rather than simple adjacency. As shown in table In a first-order autocorrelation process, tanh(μ+ is not significantly higher than for random gene pairs.Table segment effect (see Methods) was always positive in the simulated models).To validate our approach, we computed the correlation coefficients of adjacent and random gene pairs in simulated data, based on either consistent or inconsistent co-regulation. As expected, the simulated data with inconsistent co-regulation resulted in a correlation structure with only one component, whereas those with consistent coregulation showed two components. , which makes a relatively simple correlation structure plausible. Also, the data from each of their microarrays could be fit nicely with a normal mixture, which makes it possible to analyze their data on the basis of the Pearson R. As expected, most of the consistent correlation was found to be related to age group and unrelated to adjacency. Maybe more surprisingly, most of the correlation that was related to adjacency could not be accounted for by consistent co-regulation. This suggest that, at the statistical level, co-regulation of adjacent genes should be understood in terms of the mechanics of the transcription process, rather than in terms of the evolutionary origin of specific gene groups. In other words, in this particular data set, consistent correlation was very widespread, applying to 40% of all gene pairs table , half ofμ0 in table Spellman and Rubin concluded that the adjacent genes with correlated expressions were confined to certain domains, which spanned 20% of all genes. Our findings are different, in that we distinguished between two components of the correlation structure. We found a baseline correlation of 0.1  in tissue differentiation. Parisi et al.  found clμ0 was slightly positive. It is hard to imagine a biological effect that could cause μ0 to be significantly greater than zero, but it could be an artifact of the normalization: if gene g is expressed at the same level in all tissue samples, and gene h as well, they should have a correlation coefficient of zero, but since they will both yield elevated measurements in flies in which the normalization bias is positive, the observed correlation will be positive. The fact that μ0 is almost zero suggests that normalization bias is not a major problem with this data set. The fact that μ0 for random gene pairs is not generally higher in mixed age groups than in pure adult or embryo groups  gene pairs, ps table  is consi, Fukuoka found thFirst, it is not clear with respect to which anchor the physical distance should be defined. We have chosen to use the minimal distance (using the end nearest to the neighbor as an anchor), because it is not confounded by the length of the gene. However, depending on which co-regulation mechanism one has in mind, other distance measures might be more natural. This question becomes crucial if one wants to make subgroup analysis based on convergent, divergent and tandem gene pairs.Second, physical distance is a continuous variable, which means that the model must be augmented with an assumption regarding the relationship between distance and correlation.The question remains to what biological phenomena the baseline correlation between adjacent genes should be attributed. Several of the theories that have been proposed could account for it, and it is likely that several mechanisms play a role.In addition to the baseline correlation, we also found more consistent correlation between adjacent genes and inconsistent co-regulation, but this does not necessarily imply a combination of two mechanisms: one could imagine a kind of quasi-consistent mechanism, in which groups of adjacent, co-regulated genes could have boundaries everywhere, but with some boundary locations being more likely than others. Finally, it has been suggested that the correlation between the expressions of adjacent genes should be understood in terms of enhancer-promotor interaction, which weakens with distance(Dorsett). It is It is possible to analyze correlation structures in gene expression data on the basis of simple, parametric models with known mathematical properties and parameters with biological interpretations, or at least some candidate interpretations. When applied to the data from Spellman and Rubin, it appeared that the expressions of two genes can show positive correlation for either of two largely distinct reasons: because they share some confounder unrelated to adjacency , or because they are located close to each other on the chromosome. The underlying biological mechanisms remain unknown, but there appears to be a component of the correlation that depends on distance only and not of the biological function of the genes. If this is true, gene clustering algorithms might benefit from making distinction between the adjacency-related and not-adjacency-related co-regulation, for example by subtracting the effect of adjacency from the correlation of adjacent genes. Doing that would, according to our findings, lead to more reliable identifications of gene pairs with related biological functions.Ygi be the log-scale measured activity of gene g in fly i, fly i having development stage s (either "adult" or "embryo"). A natural model for Ygi would be a two-component normal mixture:First, we constructed a model for those co-regulation effects that were unrelated to the relative location of the two genes. Suppose that a gene can be active either in adults, in embryos, in both, or in neither. Let g in the other development stage should be taken into account. We therefore assumed a three-component normal mixture:Since the data were normalized so that the average logscale activity across all 89 flies was zero, the activity of gene Actually, of the 89 flies, the gene expressions in 87 of them could be nicely fitted with a three-component, normal mixture, while in two of the adult flies we identified only two components (the component of genes that were passive in adults while active in embryos vanished in those two cases).Rgh for two genes, g and h, isIf the three-component model is correct, Fisher's z-transform, which is the hyperbolic arcus tangens of the Pearson correlation coefficient μ0 to be close to zero. This model was validated by implementing it on the neighbor correlations in a shuffled data set, that is, for each gene its Pearson R with a random gene was computed. This procedure was repeated with 10 shuffled data sets, each generated by shuffling the genes in the original data set and subsequently computing the Pearson R for each pair of adjacent genes. For each shuffled data set, the parameters in the model were estimated with the EM algorithm .Second, the distribution of the correlation coefficients of the pairs of adjacent genes in the unshuffied data set were compared to the same distribution for the shuffled data sets. Doing that, we could distinguish two different contributions to the correlation; A) co-regulation effects that are unrelated to the fact that two genes are adjacent, and B) co-regulation effects related to the fact that two genes are adjacent. We also analyzed each gene's correlation with its second, third etc. (up to the tenth) neighbor, in order to get an idea of the lengths of the chromatin domains in question.Third, we applied the above two procedures to two subsets of the data, namely the data for adult flies and the data for embryos. If our assumption, that all (or at least most) correlation could be attributed to the development stage, one would expect much less correlation in those subsets. However, those subsets differ from the entire data set by the mere fact that they are smaller. Therefore, we also applied the same procedure to two random subsets of 35 and 54 flies, respectively, stratified by development stage.For those gene pairs where the start and the end of the open reading frame (ORF) was available, we defined the physical distance as the distance between the end of the ORF of the first gene and the start of the ORF of the second gene. We chose that definition because other distance definitions  would be confounded by the length of the ORFs.consistent co-regulation, in which the same segments were used for all 100 tissue samples, and one data set with inconsistent co-regulation, in which a separate segmentation was sampled for each tissue sample. If gene g belongs to segment s in sample i, the expression Ygi was given byFinally, in order to validate our method, we applied it to 6 simulated data sets, which we simulated on the basis of the two alternative hypothesis of consistent and inconsistent co-regulation. The simulated data sets contained 100 tissue samples and 1000 genes. The genes were randomly divided into either 67, 200 or 500 segments, corresponding to average segment sizes of 15, 5 and 2. This means that each gene belonged to exactly one segment. For each average segment size, we simulated one data set with Ygi = xsi + εgi     (4)xsi + and εgi were both sampled from the standard normal distribution. Notice that this model does not account for different tissue classes, so it should be compared to the results from the adults-only and embryos-only subsets. The error term εgi should be interpreted as a combination of measurement errors and biological effects that are unrelated to the segmentation.where HT did the explorative data analysis and suggested the concept of consistent co-regulation and using three-components mixtures for the correlation structure. AZ suggested using Fisher's z-transform for the correlations. HT did the literature review. Both authors contributed to the manuscript.


Plasmodium-infected mosquito bites a human, it injects the parasite—sequestered in the mosquito's salivary glands in its sporozoite stage—into the victim's bloodstream. Within hours, the sporozoites invade the liver—a critical stage for establishing infection—and spend the next few weeks asexually dividing inside liver cells, eventually releasing thousands of merozoites into the bloodstream. Merozoites quickly invade red blood cells and begin a second round of asexual proliferation. The infected cells rupture and die, releasing more parasites and toxins. The toxins cause malaria's characteristic fever and chills, and the liberated merozoites initiate another cycle of red blood cell attacks.Plasmodium, the microscopic parasite that causes malaria, passes through two hosts, two reproductive modes, four habitats, and over half-a-dozen distinct developmental stages in one lifecycle. When a An unresolved question has been how the circulating sporozoites reach the liver cells in the first place, since liver cells are separated from the bloodstream by a layer of endothelial and Kupffer cells, which form the walls of the liver capillaries. (Kupffer cells project into the bloodstream and remove contaminants.) Having identified a protein required for sporozoite migration through the capillary lining, Tomoko Ishino, Masao Yuda, and their colleagues at Mie University School of Medicine in Japan may have found an answer.Plasmodium species affect humans. P. falciparum, the most pathogenic of the human-infecting species, is closely related to avian and rodent species. One rodent species,—P. berghei—shares fundamental aspects of structure, physiology, and lifecycle with P. falciparum and so serves as a model for the human parasites. Since sporozoites must infect mosquito salivary glands before they can infect the mammalian liver, Yuda's team searched for sporozoite genes that are predicted to encode secretory or membrane proteins and are expressed only in mosquito salivary glands. Their search revealed a coding region conserved in several species of Plasmodium.Only four of the roughly 150 vertebrate-infecting Plasmodium motility and invasion, the researchers predicted this protein would also be important in migrating to or invading liver cells. They named the protein SPECT, for sporozoite microneme protein essential for cell traversal.Tracing the gene's activity through the parasite's life cycle, Yuda's team confirmed that it was expressed only in sporozoites in the mosquito salivary gland—not in the mosquito midgut, where sporozoites are produced after mosquitoes feed on the blood of an infected person. The corresponding protein was localized to micronemes, specialized secretory organelles found at the front end of malaria parasites. Because micronemes are known to play a central role in spect-disrupted mutants and observing how the altered parasites affected their hosts. spect disruption did not affect parasite proliferation in rat red blood cells or interfere with parasite development in the mosquito midgut or salivary glands, but it did have an effect on the parasite's ability to infect the liver. Rats injected with spect-disrupted parasites had significantly lower levels of liver infection than rats injected with nonmutant parasites. Since it was unclear whether the spect-disrupted mutants lost their infectivity or simply could not pass through the cell layer, the researchers inoculated human liver cells with the mutants and found that they infected the cells normally.Yuda's team tested SPECT's function by generating spect-disrupted parasites completely lost their ability to pass through cells. Since traversal of the cellular barrier between liver cells and the circulatory system is a crucial step in malarial infection, the authors conclude, SPECT and other proteins involved in shuttling sporozoites into liver cells could be effective targets for malaria treatment and prevention.Yuda's team also tested SPECT's impact on sporozoite cell-passage ability; if the mutants couldn't reach the liver cells, they couldn't infect them.


Recent technological advances in integrated circuits, wireless communications, and physiological sensing allow miniature, lightweight, ultra-low power, intelligent monitoring devices. A number of these devices can be integrated into a Wireless Body Area Network (WBAN), a new enabling technology for health monitoring.Using off-the-shelf wireless sensors we designed a prototype WBAN which features a standard ZigBee compliant radio and a common set of physiological, kinetic, and environmental sensors.We introduce a multi-tier telemedicine system and describe how we optimized our prototype WBAN implementation for computer-assisted physical rehabilitation applications and ambulatory monitoring. The system performs real-time analysis of sensors' data, provides guidance and feedback to the user, and can generate warnings based on the user's state, level of activity, and environmental conditions. In addition, all recorded information can be transferred to medical servers via the Internet and seamlessly integrated into the user's electronic medical record and research databases.WBANs promise inexpensive, unobtrusive, and unsupervised ambulatory monitoring during normal daily activities for prolonged periods of time. To make this technology ubiquitous and affordable, a number of challenging issues should be resolved, such as system design, configuration and customization, seamless integration, standardization, further utilization of common off-the-shelf components, security and privacy, and social issues. Wearable health monitoring systems integrated into a telemedicine system are novel information technology that will be able to support early detection of abnormal conditions and prevention of its serious consequences ,2. Many Important limitations for wider acceptance of the existing systems for continuous monitoring are: a) unwieldy wires between sensors and a processing unit, b) lack of system integration of individual sensors, c) interference on a wireless communication channel shared by multiple devices, and d) nonexistent support for massive data collection and knowledge discovery. Traditionally, personal medical monitoring systems, such as Holter monitors, have been used only to collect data for off-line processing. Systems with multiple sensors for physical rehabilitation feature unwieldy wires between electrodes and the monitoring system. These wires may limit the patient's activity and level of comfort and thus negatively influence the measured results. A wearable health-monitoring device using a Personal Area Network (PAN) or Body Area Network (BAN) can be integrated into a user's clothing . This syIncreased system processing power allows sophisticated real-time data processing within the confines of the wearable system. As a result, such wearable system can support biofeedback and generation of warnings. The use of biofeedback techniques has gained increased attention among researchers in the field of physical medicine and tele-rehabilitation . IntensiTypical examples of possible applications include stroke rehabilitation, physical rehabilitation after hip or knee surgeries, myocardial infarction rehabilitation, and traumatic brain injury rehabilitation. The assessment of the effectiveness of rehabilitation procedures has been limited to the laboratory setting; relatively little is known about rehabilitation in real-life situations. Miniature, wireless, wearable technology offers a tremendous opportunity to address this issue.We propose a wireless BAN composed of off-the-shelf sensor platforms with application-specific signal conditioning modules . In thisContinuous technological advances in integrated circuits, wireless communication, and sensors enable development of miniature, non-invasive physiological sensors that communicate wirelessly with a personal server and subsequently through the Internet with a remote emergency, weather forecast or medical database server; using baseline , sensor (WBAN) and environmental (emergency or weather forecast) information, algorithms may result in patient-specific recommendations. The personal server, running on a PDA or a 3 G cell phone, provides the human-computer interface and communicates with the remote server(s). Figure A WBAN can include a number of physiological sensors depending on the end-user application. Information of several sensors can be combined to generate new information such as total energy expenditure. An extensive set of physiological sensors may include the following:• an ECG (electrocardiogram) sensor for monitoring heart activity• an EMG (electromyography) sensor for monitoring muscle activity• an EEG  sensor for monitoring brain electrical activity• a blood pressure sensor• a tilt sensor for monitoring trunk position• a breathing sensor for monitoring respiration• movement sensors used to estimate user's activity• a "smart sock" sensor or a sensor equipped shoe insole used to delineate phases of individual stepsThese physiological sensors typically generate analog signals that are interfaced to standard wireless network platforms that provide computational, storage, and communication capabilities. Multiple physiological sensors can share a single wireless network node. In addition, physiological sensors can be interfaced with an intelligent sensor board that provides on-sensor processing capability and communicates with a standard wireless network platform through serial interfaces.The wireless sensor nodes should satisfy the following requirements: minimal weight, miniature form-factor, low-power operation to permit prolonged continuous monitoring, seamless integration into a WBAN, standard-based interface protocols, and patient-specific calibration, tuning, and customization. These requirements represent a challenging task, but we believe a crucial one if we want to move beyond 'stovepipe' systems in healthcare where one vendor creates all components. Only hybrid systems implemented by combining off-the-shelf, commodity hardware and software components, manufactured by different vendors promise proliferation and dramatic cost reduction.The wireless network nodes can be implemented as tiny patches or incorporated into clothes or shoes. The network nodes continuously collect and process raw information, store them locally, and send them to the personal server. Type and nature of a healthcare application will determine the frequency of relevant events . Ideally, sensors periodically transmit their status and events, therefore significantly reducing power consumption and extending battery life. When local analysis of data is inconclusive or indicates an emergency situation, the upper level in the hierarchy can issue a request to transfer raw signals to the upper levels where advanced processing and storage is available.The personal server performs the following tasks:• Initialization, configuration, and synchronization of WBAN nodes• Control and monitor operation of WBAN nodes• Collection of sensor readings from physiological sensors• Processing and integration of data from various physiological sensors providing better insight into the users state• Providing an audio and graphical user-interface that can be used to relay early warnings or guidance • Secure communication with remote healthcare provider servers in the upper level using Internet servicesThe personal server can be implemented on an off-the-shelf Internet-enabled PDA  or 3 G cell phone, or on a home personal computer. Multiple configurations are possible depending on the type of wireless network employed. For example, the personal server can communicate with individual WBAN nodes using the Zigbee wireless protocol that provides low-power network operation and supports virtually an unlimited number of network nodes. A network coordinator, attached to the personal server, can perform some of the pre-processing and synchronization tasks. Other communication scenarios are also possible. For example, the personal server running on a Bluetooth or WLAN enabled PDA can communicate with remote upper-level services through a home computer; the computer then serves as a gateway Figure .Relying on off-the-shelf mobile computing platforms is crucial, as these platforms will continue to grow in their capabilities and quality of services. The challenging tasks are to develop robust applications that provide simple and intuitive services . Total information integration will allow patients to receive directions from their healthcare providers based on their current conditions.We envision various medical services in the top level of the tiered hierarchy. A healthcare provider runs a service that automatically collects data from individual patients, integrates the data into a patient's medical record, processes them, and issues recommendations, if necessary. These recommendations are also documented in the electronic medical record. If the received data are out of range or indicate an imminent medical condition, an emergency service can be notified . The exact location of the patient can be determined based on the Internet access entry point or directly if the personal server is equipped with a GPS sensor. Medical professionals can monitor the activity of the patient and issue altered guidance based on the new information, other prior known and relevant patient data, and the patient's environment .The large amount of data collected through such services will allow quantitative analysis of various conditions and patterns. For example, suggested targets for stride and forces of hip replacement patients could be suggested according to the previous history, external temperature, time of the day, gender, and current physiological parameters . Moreover, the results could be stored in research databases that will allow researchers to quantify the contribution of each parameter to a given condition if adequate numbers of patients are studied in this manner. Again, it is important to emphasize that the proposed approach requires seamless integration of large amounts of data into a research database in order to be able to perform meaningful statistical analyses.The ActiS sensor was developed specifically for WBAN-based, wearable computer-assisted, rehabilitation applications. With this concept in mind, we integrated a one-channel bio-amplifier and three accelerometer channels with a low power microcontroller into an intelligent signal processing board that can be used as an extension of a standard wireless sensor platform. ActiS consists of a standard sensor platform, Telos, from Moteiv and a custom Intelligent Signal Processing Module – ISPM Figure . A blockThe Telos platform is an ideal fit for this application due to small footprint and open source system software support. A second generation of the Telos platform features an 8 MHz MSP430F1611 microcontroller with integrated 10 KB of RAM and 48 KB of flash memory, a USB  interface for programming and communication, and an integrated wireless ZigBee compliant radio with on-board antenna [The ISPM extends the capabilities of Telos by adding two perpendicular dual axis accelerometers  and a bio-amplifier with a signal conditioning circuit. The ISPM has its own MSP430F1232 processor for sampling and low-level data processing. This microcontroller was selected primarily for its compact size and ultra low power operation. Other features that were desirable for this design were the 10-bit ADC and the timer capture/compare registers that are used for acquisition of data from the accelerometers. The F1232 has hardware UART that is used for communications with Telos.The ISPM's two ADXL202 accelerometers cover all three axes of motion. One ADXL202 is mounted directly on the ISPM board and collects data for the X and Y axes in the same plane. The second ADXL202 is mounted on a daughter card that extends vertically from the ISPM.The user's physiological state is monitored using an on-board bio-amplifier implemented using an instrumentation amplifier with a signal conditioning circuit. The bio-amplifier could be used for electromyogram (EMG) or electrocardiogram (ECG) monitoring. The output of the signal conditioning circuit is connected to the local microcontroller as well as to the microcontroller on the Telos board via the expansion connector. The AD converter on the Telos board has a higher resolution (12 bit) than the F1232 on the ISPM (10 bit). This configuration gives flexibility of utilizing either microcontroller to process physiological signals.x and Ay. Rotations of the sensor in the vertical plane (Θ) can be estimated as Θ = arctan(Ax / Ay). A compensation for non-ideal vertical placement can be achieved using the second accelerometer (not mounted in this photo) at 90-degree angle. Instead of calculating the angular position, many systems use off-the-shelf gyroscopes to measure angular velocity for the detection of gait phases [An example application of the ActiS sensor as motion sensor on an ankle is given in Figure t phases . A typicsystem design and social issues should be resolved. If resolved successfully, WBAN systems will open a whole range of possible new applications that can significantly influence our lives.WBAN systems can capitalize on recent technological advances that have enabled new methods for studying human activity and motion, making extended activity analysis more feasible. However, before WBAN becomes a widely accepted concept, a number of challenging The development of pedometers and Micro-ElectroMechanical Systems (MEMS) accelerometers and gyroscopes show great promise in the design of wearable sensors. The main system design issues include:• types of sensors• power source• size and weight of sensors• wireless communication range and transmission characteristics of wearable sensors• sensor location and mounting• seamless system configuration• automatic uploads to the patient's electronic medical record• intuitive and simple user interfaceet al [As for sensors, accelerometers and gyroscopes offer greater sensitivity and are more applicable for monitoring of motion since they generate continuous output. Bouten et al  found thet al ,35. As eet al  systems et al . Both Laet al . Lancastet al .The majority of foot-contact pedometers are designed to count steps only. Although they have been studied for use in complex energy estimation and have even shown a high degree of accuracy for walking / running activities  they areTo be unobtrusive, the sensors must be lightweight with small form factor. The size and weight of sensors is predominantly determined by the size and weight of batteries. Requirements for extended battery life directly oppose the requirement for small form factor and low weight. This implies that sensors have to be extremely power efficient, as frequent battery changes for multiple WBAN sensors would likely hamper users' acceptance and increase the cost. In addition, low power consumption is very important as we move toward future generations of implantable sensors that would ideally be self-powered, using energy extracted from the environment.The radio communication poses the most significant energy consumption problem. Intelligent on-sensor signal processing has the potential to save power by transmitting the processed data rather than raw signals, and consequently to extend battery life. A careful trade-off between communication and computation is crucial for an optimal design. It appears that the most promising wireless standard for WBAN applications is ZigBee, as it represents an emerging wireless technology for the low-power, short-range, wireless sensors.et al use two accelerometers on the SenseWear armband [et al [et al developed a loose fitting lab coat and trousers [et al [Although the purpose of the measurement does influence sensor location, researchers seem to disagree on the ideal body location for sensors. A motion sensor attached to an ankle is the most discriminative single position for state recognition, while a combination of hip and ankle sensors discriminates the states even more . In a st armband . Lee et d [et al  placed atrousers  consistis [et al developedThe intelligent WBAN sensors should allow users to easily assemble a robust ad-hoc WBAN, depending on the user's state of health. We can imagine standard off-the-shelf sensors, manufactured by different vendors, and sold "over-the-counter" . Each seApplication-specific algorithms mostly use digital signal pre-processing combined with a variety of artificial intelligence techniques to model user's states and activity in each state. Digital signal processing include filters to resolve high and low frequency components of a signal, wavelet transform algorithms to correlate heel-strike and toe-off (steps) to angular velocity measured via gyroscopes , power sMost of the algorithms in the open literature are not executed in real-time, or require powerful computing platforms such as laptops for real-time analysis.Social issues of WBAN systems include privacy/security and legal issues. Due to communication of health-related information between sensors and servers, all communication over WBAN and Internet should be encrypted to protect user's privacy. Legal regulation will be necessary to regulate access to patient-identifiable information.The WBAN technology can be used for computer-assisted physical rehabilitation in ambulatory settings and monitoring of trends during recovery. An integrated system can synergize the information from multiple sensors, warn the user in the case of emergencies, and provide feedback during supervised recovery or normal activity. Candidate applications include post-stroke rehabilitation, orthopaedic rehabilitation (e.g. hip/knee replacement rehabilitation), and supervised recovery of cardiac patients . In the In addition, WBAN systems can be used for gait phase detection during programmable, functional electrical stimulation , analysiA wearable Wireless Body Area Network (WBAN) of physiological sensors integrated into a telemedical system holds the promise to become a key infrastructure element in remotely supervised, home-based patient rehabilitation. It has the potential to provide a better and less expensive alternative for rehabilitation healthcare and may provide benefit to patients, physicians, and society through continuous monitoring in the ambulatory setting, early detection of abnormal conditions, supervised rehabilitation, and potential knowledge discovery through data mining of all gathered information.Continuous monitoring with early detection likely has the potential to provide patients with an increased level of confidence, which in turn may improve quality of life. In addition, ambulatory monitoring will allow patients to engage in normal activities of daily life, rather than staying at home or close to specialized medical services. Last but not least, inclusion of continuous monitoring data into medical databases will allow integrated analysis of all data to optimize individualized care and provide knowledge discovery through integrated data mining. Indeed, with the current technological trend toward integration of processors and wireless interfaces, we will soon have coin-sized intelligent sensors. They will be applied as skin patches, seamlessly integrated into a personal monitoring system, and worn for extended periods of time.


The recently created professional science master's degree may be the answer to the increasing need for science-savvy employees in the business world Sloan Foundation to cultivate science managers. Sloan's ultimate goal is to make science careers more attractive to talented young people like Herzer, a 2003 PSM graduate in Applied Biosciences from the University of Arizona, who believes his PSM makes him more marketable to science-based businesses. “I am at the true junction of science and business,” he says.Fortunately for Herzer, the business of science is booming. Jobs for scientists and engineers grew four times faster than the United States national average since 1980, and should outpace the market until at least 2010. Surprisingly to many academics, most of these jobs are in industry. In 1999, the last year with complete data, two out of three employed science and engineering (S&E) graduates worked in industry, including the great majority of bachelor's and master's degree holders, and 40% of doctorates. In other words, industry, not academe, now drives American S&E employment, and will for the near future.Like academia, industry needs scientifically literate personnel; unlike academia, industry wants employees with business savvy as well. However, in the past, graduate students received either science or business instruction, not both. “Industry simply hired regular master's-degreed people, or MBAs, or more likely PhDs, and just expected them to learn their weaknesses on the job,” says Eleanor L. Babco, Executive Director of the Commission on Professionals in Science and Technology, a nonprofit corporation with funding from the Sloan Foundation to assess PSM graduates.For science-based businesses, then, the American S&E doctorate—viewed by many as the worldwide gold standard for science education—is too specialized for their needs see . But a mThe length of time to obtain a biological science doctorate has increased……The number of postdocs and part-time faculty in the biological sciences is increasing……And the proportion of doctorates in academia is decreasing…http://www.nsf.gov/sbe/srs/seind04/] and the National Science Foundation Science Resources Statistics Division's “1995 and 2001 Survey of Doctorate Recipients” [http://www.nsf.gov/sbe/srs/infbrief/nsf04328/table1.xls].) program.Like all graduate programs, PSMs offer advanced coursework in a (science or math) specialization, usually in an emerging or hybrid field such as bioinformatics. Most PSMs also provide business courses—including finance, project management, regulatory affairs, and intellectual property law—and information technology classes as well. PSMs are “industry relevant” by design, with external advisory committees populated by local business leaders, weekly colloquia led by corporate representatives, special arrangements for employed students, and industry internships or final projects exploring realistic business scenarios see .Only programs meeting most of the following requirements may earn the official moniker “professional science master's.”Two years of science or math graduate-level coursework, taught by regular faculty, characterized by interdisciplinary studies and a focus on informaticsTraining in business fundamentals—such as finance, marketing, project management, communication, and team building—and exposure to industry professionalsFinal project reflecting a realistic workplace issue and/or industry internshipAdvisory board of industry professionalsTargeted recruitment and admissions separate from other degree programsCommitment to tracking graduates through first five yearsLong-term sustainability http://www.sciencemasters.com/affiliation.html.)Wall Street Journal, “Harvard tries to create leadership in industry, academics and government, and our philosophy is we don't think that with a master's degree people can fill that role very easily.”Although most scientific careers demand a graduate degree, a professional master's in many hard sciences still encounters entrenched academic opposition. According to Lee-Jen Wei, then acting chair of the Department of Biostatistics in the Harvard School of Public Health, quoted in the The government appears to agree with this view. While most doctoral candidates receive federal funds for tuition and other expenses, there is little money for master's students, who disproportionately end up in industry regardless of specialization. PSM students are especially affected by this problem because “interdisciplinary” equals “expensive.” Similarly, “interdisciplinary” can also mean “hard to find”—companies with targeted recruitment often miss PSM students, who are not “in” any particular department—and “confusing”—differences in these new, still somewhat vaguely defined programs can make hiring comparisons difficult. But perhaps the most conspicuous drawback to PSMs is their newness, and resulting obscurity: almost half of graduates say they are “not sure” employers will value their PSM, or the unique skill set it affords.Professionalizing Graduate Education: The Master's Degree in the Marketplace, argues that converging market forces will lead to the success of the professional master's degree in science. These forces include: rapid technological change; the rise of alternative learning channels such as online and distance education, corporate universities, and hi-tech certification programs; the proliferation of degrees in general, and in multidisciplinary fields specifically; and a fundamental societal shift away from public service and toward entrepreneurship, profitability, and competition.Still, many observers of higher education support the PSM concept. Judith Glazer-Raymo, author of the forthcoming book Kenneth R. Smith, former dean of the Eller College of Business and Public Administration at the University of Arizona, and others make the case that PSMs may protect students' careers from outsourcing to foreign countries. The American S&E labor pool is shrinking, and industry has already responded by transferring much of its research and development overseas; however, companies are mostly moving lab scientists, not strategic analysts. Cross-training in both science and business could thus provide an edge for domestic workers in the near-term employment environment; in fact, PSM programs have a higher proportion of US citizens and residents than S&E doctoral programs.Further, in its 2003 report, the National Science Board urged the government to better align S&E graduate education with “expected national skill needs,” including “interdisciplinary skills.” The report also recommended federal funding for a wider range of educational options and more attention on the real economic concerns of students—code words for support of professional master's degree initiatives. In the same vein, top universities now advocate “interconnections” between their professional schools and traditional departments, as a way of strengthening the overall academic mission, and many countries are sponsoring initiatives to stimulate university–industry links, to maximize marketing of technological innovations.For advocates, then, the PSM both advances the cause of science education reform and addresses changing employment conditions with one big idea: reinvention of the two-year graduate credential for an entrepreneurial age. Herzer, for one, now a technology development representative at the Scripps Research Institute, has staked his future on the potential of professional master's degrees. “Scientists rarely understand business dealings, and business personnel rarely comprehend scientific discoveries,” he says. “The overlay of the two is crucial for any successful business transaction of scientific origins.”


EIF2B2, 4 and 5.Premature Ovarian Failure (POF), defined as the development of hypergonadotropic amenorrhea before the age of 40 years, occurs in about 1% of all women. Other than karyotype abnormalities, very few genes are known to be associated with this ovarian dysfunction. Recently, in seven patients who presented with POF and white matter abnormalities on MRI (ovarioleukodystrophy) eight mutationswere found in EIF2B genes in POF, we screened 93 patients with POF who did not have identified leukodystrophy or neurological symptoms. We evaluated these eight mutations and two additional mutations that had been found in patients with milder forms of eIF2B-related disorders. We used restriction enzymes and direct sequencing.To further test the involvement of known mutations of EIF2B genes, either homozygous or heterozygous, were identified in our 93 patients with pure 46,XX POF. The upper 95 % confidence limit of the proportion 0/93 is 3.2%.None of the known mutations in We conclude that eIF2B mutations, already described in cases of POF associated with white matter abnormalities, are an uncommon cause of pure spontaneous premature ovarian failure. Premature Ovarian Failure (POF) can present as a primary or secondary amenorrhea and is associated with elevated gonadotropins before 40 years of age. POF affects 1% of all women and occurs in 0.1% before the age of 30 years . While cEIF2B genes  were reportedly involved in seven patients who presented with POF and white matter abnormalities on MRI (ovarioleukodystrophy) [spectrum can range from a rapid course leading to death in severe congenital forms to asymptomatic MRI findings in adult patients [EIF2B genes.Recently three of the five strophy) . These gpatients ,11. Ovarpatients . TherefoEIF2B mutations in 93 unrelated and well-characterized women with POF. An institutional review board approved the study and all participants gave a written informed consent. Referring physicians made the diagnosis of premature ovarian failure based on the following criteria: development of at least 4 months of amenorrhea before age 40 associated with two serum FSH levels in the menopausal range. Women with premature ovarian failure as a result of surgery, radiation, chemotherapy, or known karyotype abnormalities were not included in the study. There were 6 Asians, 12 Blacks, 4 Hispanics and 71 Caucasians. The median age at the onset of menstrual irregularity was 24.5 years (range 13 to 39). Eighteen women had a family history of POF. All women underwent a history and physical examination and laboratory screening to confirm the diagnosis of POF and all had a normal karyotype. None of the women had evidence of a neurological disorder.In the current study, we evaluated the presence of Genomic DNA was extracted from peripheral blood using standard procedures.EIF2B2, 4 and 5 which contain mutations found in POF patients or in milder forms of eIF2B-related disorders were amplified by the polymerase chain reaction (PCR) as previously described  mutation in the escribed .EIF2B4) described in milder forms of eIF2B-related disorders were not found in this series of 93 patients with POF. The upper 95 % confidence limit of the proportion 0/93 is 3.2%.None of the eight mutations already described in ovarioleukodystrophy were detected in our 93 patients with pure 46,XX POF, neither in a homozygous nor in a heterozygous state. In addition, the mutations C728T and C1120T , 2 to 5 years (group 2) and > 5 years (group 3) . Group 3In these six eIF2B-mutated families, neurological symptoms with abnormalities of the cerebral white matter on MRI were associated with primary or secondary amenorrhea due to POF . A correEIF2B mutations. In the present study, we tested for EIF2B mutations a series of 93 patients with pure, karyotypically normal POF without identified signs of cerebral dysfunction.The mutated eIF2B may be responsible for both increased apoptosis of ovarian follicles leading to POF, and a defect in glial cell development causing abnormal formation of white matter structures. In ovarioleukodystrophy, a phase of amenorrhea without neurological symptoms can be observed, suggesting that an apparently isolated case of POF might be due to EIF2B5 gene) and A638G (EIF2B2 gene) are found in 71% of families with late onset forms of eIF2B-related disorders (group 3) [EIF2B mutations in women with POF who have no apparent neurological signs.In eIF2B-related disorders, a correlation exists between genotype and disease onset . The mutgroup 3) . In ovarEIF2B mutations is not clinically indicated.For patients presenting with POF without neurological signs or MRI abnormalities, the routine screening of the The author(s) declare that they have no competing interests.AF and FGB carried out the molecular genetics studies, including enzyme restrictions (AF) and sequencing (AF and FGB). AF drafted and conceived of the study. RS participated in the coordination of the study. VHV, VKB, LMN recruited and evaluated the patients, collected DNA samples, participated in the design and coordination of the study, and helped in drafting the manuscript. OBT conceived of the study, and participated in its design and coordination. All authors participated in the writing of the manuscript and have read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


Drosophila melanogaster genes involved in hybrid incompatibility have been identified but none has yet been shown to have functionally diverged in accordance with the Dobzhansky-Muller model. By introducing transgenic copies of the X-linked Hybrid male rescue (Hmr) gene into D. melanogaster from its sibling species D. simulans and D. mauritiana, we demonstrate that Hmr has functionally diverged to cause F1 hybrid incompatibility between these species. Consistent with the Dobzhansky-Muller model, we find that Hmr has diverged extensively in the D. melanogaster lineage, but we also find extensive divergence in the sibling-species lineage. Together, these findings implicate over 13% of the amino acids encoded by Hmr as candidates for causing hybrid incompatibility. The exceptional level of divergence at Hmr cannot be explained by neutral processes because we use phylogenetic methods and population genetic analyses to show that the elevated amino-acid divergence in both lineages is due to positive selection in the distant past—at least one million generations ago. Our findings suggest that multiple substitutions driven by natural selection may be a general phenomenon required to generate hybrid incompatibility alleles.Interspecific hybrid lethality and sterility are a consequence of divergent evolution between species and serve to maintain the discrete identities of species. The evolution of hybrid incompatibilities has been described in widely accepted models by Dobzhansky and Muller where lineage-specific functional divergence is the essential characteristic of hybrid incompatibility genes. Experimentally tractable models are required to identify and test candidate hybrid incompatibility genes. Several  Transgenic experiments show that the HMR gene has functionally diverged in Drosophila melanogaster and its sibling species and causes the death of hybrid offspring in interspecific crosses A from one species causes the incompatibility while the ancestral allele a from the other species does not.Reproductive isolation is the most commonly used criterion to define species. Hybrid incompatibilities (HIs) such as hybrid sterility and lethality are widely observed examples of reproductive isolation. The Dobzhansky-Muller (D-M) model explains how the genes causing deleterious phenotypes in hybrids can evolve  Figure A. The moA (or X) but not allele a (or x) will affect the HI phenotype. For example, increasing the dosage or activity of allele A should decrease hybrid fitness, while identical manipulations of allele a should not. In contrast, downstream effector genes such as d1 in Drosophila melanogaster.This model makes clear and testable genetic predictions, namely, that experimental manipulation of allele If the genetic changes that cause HI are rare, then genes that have undergone extensive divergence may be more likely to cause HI simply because there is more chance that they have experienced rare HI-causing mutations. But the D-M model itself offers no suggestions about the mode of evolution that leads to this divergence. One possibility is that HI genes accumulate genetic changes over time as they evolve neutrally. Alternatively, many models have suggested that speciation may be driven by molecules that are undergoing natural selection, for example, under the pressure of ecological divergence  or sexuaD. melanogaster can hybridize with three closely related species that we refer to collectively as its sibling species: D. simulans, D. mauritiana, and D. sechellia. Crosses between D. melanogaster females and sibling-species males result in invariantly lethal hybrid sons and temperature-dependent lethal hybrid daughters  gene has a major effect on the fitness of hybrids from this cross. This point is most strikingly demonstrated by the fact that D. melanogaster Hmr loss-of-function mutations such as 1Hmr suppress the lethality of both hybrid males and females (In(1)AB rescue mutation  from the sibling species. Hmr gene, which encodes a predicted DNA-binding protein similar to the ADF and MYB family of transcriptional regulators, and proposed that D. melanogaster HI may be caused by transcriptional misregulation.aughters . The Hyb females , as doesmutation . Increasiability . These sHmr is that all genetic manipulations were done only with the D. melanogaster allele. These studies therefore cannot determine whether Hmr is an HI gene as modeled by Dobzhansky and Muller  sites, a remarkable finding considering that Hmr encodes a predicted protein over 1,400 amino acids in length. Two other Drosophila genes that are involved in HI show elevated divergence that appears to have evolved under positive selection, but the divergence is confined to a small region of each gene AB mutations AB mutations rescuing hybrids by reducing or eliminating +Hmr activity. The transgenic copies of +D. melanogaster Hmr thus suppress rescue by increasing +Hmr activity in hybrids. We therefore asked whether or not sibling-species +Hmr has the same activity. We reasoned that if Hmr is functionally diverged between the species, as in +Hmr would not have the property of suppressing hybrid rescue. On the other hand, if Hmr is not functionally diverged, as in +D. melanogaster Hmr transgenes and suppress hybrid rescue.In the absence of available nogaster . It has utations . Those e+D. melanogaster Hmr transgenic construct and that exceed the minimal +Hmr region previously defined AB in D. melanogaster/D. simulans hybrids AB-dependent rescue of hybrid female sterility AB,w/X females heterozygous for the insertion +t8.6}2-4P{Dsim\Hmr averaged 10.0 ± 8.9 eggs (n = 41) while their non-transgene-carrying sisters averaged 8.0 ± 12.4 eggs (n = 28). Using a second, independent transgenic line, we found that sim,wIn(1)AB,w/X females heterozygous for the insertion +t8.6}4-1P{Dsim\Hmr had 12.2 ± 13.7 eggs (n = 24) while their non-transgene-carrying sisters had 17.7 ± 17.3 eggs (n = 31).To maximize the chances that the sibling-species transgenic lines would function normally, we made constructs that are similar to the largest ined see A. We ass hybrids . In all terility . In contansgenes , we foun+Hmr transgenes are expressed  site N)(D between D. melanogaster and its three sibling species (Hmr between D. melanogaster and D. simulans and compared it to compiled datasets containing over 250 genes from these two species (Hmr has one of the highest levels of nonsynonymous divergence (0.089); only four other non–Accessory gland protein (Acp) genes have a higher nonsynonymous divergence, and two of these include expressed sequence tag comparisons less than 350 bases in length. The remaining two loci, both X-linked, are mei-218 and Odysseus. In contrast, the average divergence per synonymous site (Ds) of 0.110 for Hmr is lower than the mean value of 0.125 for this dataset (again excluding Accessory gland protein genes).It has been shown previously that  species . To unde species . PairwisHmr raises a number of important questions: (1) Is divergence high on the lineage of either D. melanogaster or its sibling species, or both? (2) Was the divergence on either or both lineages caused by positive selection consistent with the time scale of speciation? (3) Can we identify specific regions and codons subject to positive selection on these lineages, and how does this compare to the few other known candidate speciation genes? (4) Is divergence on one or both lineages potentially responsible for causing the HI phenotype of Hmr?The exceptional divergence of Hmr began to diverge rapidly after the D. melanogaster–sibling-species divergence by isolating and assembling an outgroup Hmr orthologous sequence from the D. melanogaster subgroup species D. erecta, which is estimated to have diverged from D. melanogaster between 6 and 15 million years ago , then one would expect that the ratio of nonsynonymous to synonymous divergence between species would be similar to the ratio of nonsynonymous to synonymous polymorphisms found within these species (π and synonymous S) and D. simulans  relative to other loci . This significant variation across the locus can be further refined by examining the ratio of nonsynonymous to synonymous variation across the locus and is visually displayed for D. melanogaster and D. simulans using sliding window analyses in ND/SD ratios and low Nπ/Sπ ratios, in other words, much more amino-acid divergence than polymorphism. These contrasting ratios do not suggest that regional variation in substitution rates among the two classes of sites is due solely to mutation (and drift) but rather suggest that selection is contributing to the divergence pattern across the gene. Although this sliding window plot is highly suggestive of selection, we wished to obtain independent and statistically supported evidence for regional selection. We therefore tested rates of divergence relative to polymorphism for each exon in a manner similar to the McDonald-Kreitman test (MK test) described in the previous section. The data were partitioned a priori using each exon as a unit (rather than picking an arbitrary window size). Given the size of the sampled region, there is considerable power to address regional variation. These MK tests for each exon revealed that the fourth and fifth exons of Hmr appear to contribute the most to the overall MK test . It is interesting to note that these regions are not homologous to any other proteins known in D. melanogaster (or in any other species), including other MADF domain–containing proteins. We conclude that multiple regions of Hmr show strong evidence for positive selection.Polymorphism and divergence vary substantially for the separate exons and the DNA binding domains. In contrast to the neutral expectation, we found that there is highly significant heterogeneity in the ratio of polymorphism to divergence across the five exons of Hmr locus appear to have evolved rapidly since D. melanogaster and the sibling species diverged, relative to both synonymous coding sites and introns. The ratios of divergence to polymorphism between D. melanogaster and D. simulans for the 5′ and 3′ flanking regions combined (including the UTRs) are significantly different from those of the coding and intron regions  . These observations suggest that adaptive fixations have occurred at a number of noncoding sites.We also observed that the flanking regions of the  0.0001) , but thiHmr might offer a unique opportunity to apply these approaches to identify codons that exhibit positive selection and that could be contributing to HI. We identified 25 amino-acid positions that may have diverged due to selection, 20 of which fit possible models for the evolution of the D. melanogaster–sibling-species incompatibility . Based on these values we expected to find approximately three to four shared polymorphic sites for Hmr among the 54 segregating sites in D. melanogaster and 94 segregating sites in D. simulans, but instead found zero. Although the expectation of three to four shared polymorphisms is not a strong test, these findings are consistent with Hmr alleles not having been exchanged between these taxa since at or near the time of species divergence and/or with selection events having occurred throughout the Hmr gene and having removed recurrent mutations.Loci directly involved in reproductive isolation may reflect the true species history of divergence more accurately than a “random” locus sampled from the genome, because these loci cease exchanging alleles among species earlier than other loci . Both reears ago , and manears ago . Howeverears ago , we founHmr is due to positive selection. These findings raise the critical question of whether selection events occurred recently or at the time of speciation. The number of years separating D. melanogaster and the sibling-species lineage makes this a challenging task. We therefore concentrated on the more tractable problem of looking for evidence to exclude the possibility that Hmr divergence reflects the action of contemporary or recent selection. In other words, while it may not be possible to prove that selection occurred at Hmr at the time of speciation, it is nevertheless important to demonstrate that available data are, at the least, not inconsistent with more ancient selective events. Examination of the frequency spectrum of polymorphism within both D. melanogaster and D. simulans showed no evidence of recent sweeps, as the distribution of per-site heterozygosities did not differ from expectations of neutrality . However, the power for detecting a selective sweep with this method is not high (75%) even if the event occurred in the very recent past, and diminishes to 25% as early as 0.8N generations ago (where N= population size)  is clearly not consistent with the polymorphism data, whereas adaptive mutations occurring more than N generations ago are consistent with the polymorphism data, and selective sweeps more than 1.5N generations ago are possible  . Becausehappened . To estipossible . Similarvolution  but haveD. simulans or +D. mauritiana Hmr have no effect on hybrid fitness, in contrast to the strong deleterious effects previously observed with +.D. melanogaster Hmr These transgenic experiments demonstrate that Hmr has functionally diverged between D. melanogaster and its sibling species and thus meets the experimental criteria defined by the D-M model. To our knowledge this is the first demonstration of functional divergence for a D. melanogaster HI gene. Although transgenic technologies have limitations—for example, genes with large and complex regulatory regions may not function correctly when transformed—they have the clear advantage of providing unambiguous analyses of single genes. Transgenics was previously used  is a gene duplication that is present in X. maculatus but appears to be absent from X. helleri). Functional divergence can also be inferred in other HI systems where multigene regions are transferred between species by repeated backcrossing. This approach has been used to show that Odysseus (Ods) has functionally diverged: X-chromosome introgressions from D. mauritiana into D. simulans that contain the D. mauritiana Ods region are sterile, while related introgressions that lack D. mauritiana Ods remain fertile (We have shown here that transgenes carrying sly used  to show Weis and ). Functi fertile .Hmr and Ods, as well as Nup96, which has recently been implicated in causing lethality in D. melanogaster/D. simulans male hybrids ((Ods and Nup96) or most (Hmr) of their coding regions. If generally true, this finding may reflect the fact that a high rate of substitution is required in order to generate a rare HI-causing mutation. Alternatively, it may be an indication that the combined effects of multiple substitutions are required to generate an HI allele. Our analysis of Hmr suggests two additional characteristics of HI genes. One is that while the D-M model requires divergence along only one lineage in order to generate HI, we have strong statistical support for accelerated divergence of Hmr along both lineages since D. melanogaster and the sibling species split.Both  hybrids , show hiHmr that are candidates for causing the incompatibility. The simple model shown in melHmr and sib,A implies that any codons in melHmr that have diverged from the ancestral allele could potentially be causing the incompatibility; these codons could also diverge in the sibling-species alleles as long as they remain different from the D. melanogaster allele. There are 137 amino acids in Hmr (plus two sites of D. melanogaster–specific insertions) that fit these criteria. However, considering the extensive divergence we have detected at Hmr along both lineages, a more realistic model is shown in Hmr and gene A go through multiple changes in both D. melanogaster and the sibling species. This model raises a possibility described by sibA may become incompatible with anc,Hmr which means that melHmr will also be incompatible if it retains the interacting residues present in anc.Hmr Since sibHmr must be compatible with sib,A the candidate codons are those where sibHmr differs from ancHmr and mel,Hmr and melHmr remains identical to anc.Hmr There are 49 amino acids that fit this model, all of which are different from those identified in the previous model. Distinguishing between these models may be possible by using our transgene assays on +Hmr constructs containing site-directed changes at candidate amino acids.One consequence of this extensive divergence is that it becomes unexpectedly complicated to identify the divergent codons in Hmr as well as Ods and Nup96. Adaptive fixations and reproductive isolation at Hmr have clearly shaped the pattern of polymorphism relative to divergence and swept away any shared polymorphisms that may have been present. This characteristic raises the general question of what forms of selection are responsible and whether genes involved in certain traits or phenotypes that are under strong directional selection may preferentially contribute to HI. In allopatric models, where speciation occurs between two populations in geographical isolation, HI between species is strictly a secondary consequence of divergence of HI genes that has occurred within each species. The target of selection must therefore be sought by looking at gene function within species. We do not yet know the function of 1Hmr; Hmr mutants are viable and fertile, but this allele is clearly hypomorphic  ancestor, respectively.The subscripts RT-PCR was performed from total RNA as described in Hmr constructs were derived from D. simulans and D. mauritiana lambda genomic libraries  to make the p94 plasmid. An approximately 9.4-kb NotI/XbaI fragment of the p94 insert was cloned into the corresponding sites of pCasper4. End sequencing of the insert demonstrated that this construct extends from approximately 2.8 kb 5′ of Hmr to approximately 1.8 kb 3′ of Hmr. This construct also contains the complete predicted coding regions of Rab9D and CG2124. The formal designation of this construct is +mc Dmau\Rab9D+t9.4 Dmau\Hmr+t9.4 Dmau\CG2124+t9.4=Dmau\Hmr+t9.4}.P{wA D. erecta Hmr was isolated by screening a gridded fosmid library  with D. melanogaster Hmr cDNA clone RE54143 as a probe. An approximately 8-kb XbaI fragment was subcloned from fosmid 11D-19 into pBSII KS(+) and sequenced using a GPS-1 Genome Priming System  and custom sequencing primers.Hmr ortholog from the more distant D. pseudoobscura, but an unalignable repetitive region in the second exon made unambiguous calculations of amino-acid and silent divergence impossible.We also identified an D. melanogaster, the ortholog of CG2124 is located 5′ of Hmr in opposite orientation in both D. erecta and D. pseudoobscura, demonstrating that we have correctly identified the ortholog of Hmr in both of these species.As is found in D. melanogaster Hmr alleles were sequenced from a collection of iso-X-chromosome stocks derived from isofemale lines collected in Zimbabwe . We also included in our analyses four D. melanogaster alleles and one each from the three sibling species reported previously , Hmr coding region from D. mauritiana contained in plasmid p94 (AY573924), Hmr coding region from D. simulans contained in plasmid p92 (AY568391), D. melanogaster Hmr alleles sequenced from iso-X-chromosome stocks derived from isofemale lines collected in Zimbabwe (AY568380–AY568389), and D. simulans Hmr alleles sequenced from highly inbred isofemale lines collected in Wolfskill, California, United States (AY568392–AY568396).The GenBank accession numbers of the genes discussed in this paper are


Peach is being developed as a model organism for Rosaceae, an economically important family that includes fruits and ornamental plants such as apple, pear, strawberry, cherry, almond and rose. The genomics and genetics data of peach can play a significant role in the gene discovery and the genetic understanding of related species. The effective utilization of these peach resources, however, requires the development of an integrated and centralized database with associated analysis tools.The Genome Database for Rosaceae (GDR) is a curated and integrated web-based relational database. GDR contains comprehensive data of the genetically anchored peach physical map, an annotated peach EST database, Rosaceae maps and markers and all publicly available Rosaceae sequences. Annotations of ESTs include contig assembly, putative function, simple sequence repeats, and anchored position to the peach physical map where applicable. Our integrated map viewer provides graphical interface to the genetic, transcriptome and physical mapping information. ESTs, BACs and markers can be queried by various categories and the search result sites are linked to the integrated map viewer or to the WebFPC physical map sites. In addition to browsing and querying the database, users can compare their sequences with the annotated GDR sequences via a dedicated sequence similarity server running either the BLAST or FASTA algorithm. To demonstrate the utility of the integrated and fully annotated database and analysis tools, we describe a case study where we anchored Rosaceae sequences to the peach physical and genetic map by sequence similarity..The GDR has been initiated to meet the major deficiency in Rosaceae genomics and genetics research, namely a centralized web database and bioinformatics tools for data storage, analysis and exchange. GDR can be accessed at  Malus), pear (Pyrus), raspberries/blackberries (Rubus), strawberries (Fragaria), and stone fruits (Prunus) such as peach/nectarine, apricot, plum, cherry and almond [Arabidopsis and other characteristics: a relatively short juvenile period (2–3 yrs) and extensive genetics and genomics resources such as molecular marker maps, interesting mutants and clone library resources [In the United States and temperate regions throughout the world, the family Rosaceae ranks third in economic importance. The most important fruit producing crops include apple  has been initiated. The goals of the GDR are (1) to develop an organized and integrated web resource for peach genomics data to facilitate the gene discovery in other member species by a comparative mapping approach, (2) to collect and integrate all Rosaceae genomics data, (3) to develop online tools and resources for the Rosaceae community. In this paper, we describe the structure and content of the database, we review the database access utility and tools and we report a case study where we mapped publicly available Rosaceae sequences to the peach physical and genetic maps by sequence similarity.The volume and the complexity of the data being produced by these peach genomics projects, in addition to the rapidly accumulating genomics and genetics data for other important rosaceous species, necessitate the development of a properly curated and integrated scientific database. Such a database will help scientists to efficiently access, analyze, integrate and apply the data to their own research in a timely manner. RoseDB, which focused on apple genetics and cherry genomics, has been decommissioned and now only exists as a mirror site at INRA. To meet the major need for a centralized and integrated web-database for genomics and genetics research in Rosaceae, the Genome Database for Rosaceae  and the singletons that have either unique protein matches or no known significant matches. Peach ESTs were further annotated by Gene Ontology (GO) assignment based on the single "best hit" match against the SWISS-PROT database. Of the 1552 sequences from the putative peach unigene set that had matches with the SWISS-PROT database, 1439 could tentatively be assigned GO classifications. Additional sequence annotation includes computational analysis for simple sequence repeats (SSR) and open reading frame (ORF) on both the filtered clone library and the contig library. SSR analysis was preformed using a modified version (CUGISSR) of a Perl script SSRIT [. FLIP is a UNIX C program that finds/translates ORFs (open reading frames) in sequences. Using the FLIP output, CUGISSR selects the longest ORF as the putative coding region and reports the location of SSRs in relation to the putative coding region.The peach and almond ESTs are being processed at the Clemson University Genomics Institute (CUGI) utilizing publicly available software, integrated in a fully automated in-house developed script (CUGIEST). The processing occurs in three stages: trace file processing to identify a filtered, high quality clone library, assembly of high quality sequences to produce longer transcripts and reduce redundancy, and sequence annotation. Annotation consists of pairwise comparison of both the filtered clone library and the EST contig consensus sequences against the GenBank nr protein database using the fastx3.4 algorithm . The tenpt SSRIT  with parpt SSRIT  program -9) following a monthly homology search of GenBank nr protein database using the fastx34 algorithm.In addition to the peach and almond ESTs processed by CUGI, all the publicly available Rosaceae EST data are daily downloaded from GenBank dbEST and annotated with the top ten most significant matches . The SVG viewer plug-in can be freely downloaded from the Adobe Website  and the system requirements can be found at their website . Our map viewer program accesses our underlying relational database to dynamically generate an integrated genetic and transcript map with a direct link to the WebFPC physical map from each marker.Web interfaces for database query and the query result pages are mostly developed using Java Server Page (JSP). JSPs are more efficient, easier to use, more powerful and more portable than traditional CGI and many alternative CGI . BAC conThe GDR website is composed of general information pages, database query/browse interfaces and other tools such as map viewer and sequence similarity server. The GDR web pages are extensively linked such that users can easily access the data of interest regardless of the navigation starting point. For example, the EST detail pages have links to the BAC detail pages, marker detail pages or map viewer for the ESTs that are anchored to BACs, markers or maps. Similarly, the BAC detail pages have links to EST detail page, marker detail page, WebFPC and map viewer. Users can also access the data detail pages from the map viewer or WebFPC/WebChrom. Instead of displaying the entire EST or BAC data in one page, we used the right hand side navigation bars to help users find specific information easily and quickly. A general GDR navigation tool bar is also included in each page to help provide a more user-friendly interface.The generic search site allows users to select data types such as EST, BAC and Marker, and search by name. Users can also follow the link to perform more detailed search for each data type. In the EST search site, users can search either the CUGI peach EST database or Rosaceae EST database downloaded from GenBank. ESTs can be searched by their name(s) and annotation features such as whether the EST belongs to a contig, is a unigene, is used as a probe and has SSRs, or any combination thereof. The EST details page, instead of displaying all the details in one page, initially displays the clone information and the sequence with a side bar containing links to library detail, assembly/unigene information, sequence homology, SSR information, Map position and anchored BACs. Each page linked from this side bar has the same side bar for easy navigation between the features. The Sequence homology page shows the most significant matches (EXP < 1e-9) in the Genbank nr protein database from the fastx sequence similarity search. The SSR information page shows the sequence along with the computationally derived SSRs to help users in the primer design for SSR marker development. The longest putative open reading frame (ORF) is also marked in color in the sequence along with the SSRs. SSRs in the non-coding region tend to be more polymorphic and those in the coding region tend to be more transferable among species, so the information of SSR position in a gene structure will be useful for marker development. The Map position page allows users to view the ESTs' map position using our Map Viewer. Users can retrieve the anchored BAC clones for the EST of interest and all the other ESTs and markers that hybridized to the same BAC through the anchored BACs page. The assembly/unigene information page displays the assembly results which include the contig name and the unigene clone that best represents the contig. The contig name is linked to a contig page that displays the contig sequence with a side bar containing links to the comprising ESTs, sequence homology and SSR information. As ESTs with no match to the GenBank proteins or with no SSRs can still assemble into contigs with a match or SSRs, users may get further annotation results of their ESTs of interest by visiting the contig detail site. In addition, contigs may have longer sequences surrounding the SSRs, allowing more flexibility in the primer design for marker development.In the BAC search site, users can search BACs by name or by probe specifications used for BAC hybridization. The search results site and the linked sites provide users with all the data about the BAC, such as the BAC contigs that the BAC belongs to, other BACs in the contig, probes that hybridized to the BACs, the detailed data about the probes, and link to the WebFPC physical map. Markers can be searched by name or features such as map name, type, and source organism. Similarly with the BAC search result sites, the maker search results sites leads users to pages with the marker information, anchored BACs, other markers and ESTs hybridized to the same BACs and link to the Map Viewer and the WebFPC physical map.Prunus map and each linkage group has a link to a page where the developing contigs are located by the linkage group. Each contig has a direct link to WebFPC in which the individual BAC clones are displayed  and various search parameters. Users can upload batch sequence files and the parsed results from the search are formatted in a spread sheet and sent to users by email. When the query sequence has a match to the annotated GDR sequences, users can retrieve all the information such as putative function, SSRs, and the anchored map positions via a hyperlink in the excel spreadsheet. Our sequence similarity server, specifically designed for Rosaceae researchers, will help users utilize the developing peach resources in the studies of other Rosaceae species. For example, as described in the case study below, sequences derived from other Rosaceae species could be immediately anchored to the various Rosaceae maps and the physical map when the query sequences show significant similarity to the mapped peach ESTs.Prunus sequences such as apricot (Prunus armeniaca), almond (Prunus dulcis) and sour cherry (Prunus cerasus). This was expected as peach also belongs to the genera Prunus. The 259 query/match pairs consisted of 209 Rosaceae sequences and 61 mapped peach ESTs. The matching of multiple Rosaceae sequences to single peach sequences was expected since the Rosaceae sequences were not assembled and therefore potentially contains multiple sequences representing the same gene. The 209 Rosaceae sequences were anchored to 38 different loci in four different Rosaceae maps. The number of sequences from each Rosaceae species that anchored to each map is shown in Figure .We report here a case study illustrating the utility of our sequence similarity server and other integrated GDR web resources. In this study, we performed a sequence similarity search using the FASTA algorithm with the non-peach Rosaceae sequences against the mapped peach ESTs to annotate Rosaceae sequences with map positions. A fasta formatted file with a total of 16258 publicly available non peach Rosaceae sequences was uploaded to the GDR FASTA server. We selected the mapped peach database and used the default parameters. The search results returned from the server are formatted in a spread sheet for easy browsing and the match names are hyperlinked to both GenBank and the GDR web site (Figure Prunus genomics projects. Data to be added in the near future include apple ESTs, strawberry ESTs, rose ESTs and apricot map/marker data from collaborators. When the genomics projects from Prunus are finished, we will host 10–15,000 unique ESTs from a variety of vegetative and reproductive tissues of peach and almond, the complete peach physical map with anchored genetic markers and unique ESTs. In addition to adding new data, future development efforts will focus on improving the tools and functionality of the web interface such as an advanced search site with options for search/display categories, full sequence processing facilities for Rosaceae researchers, a newsgroup for the Rosaceae community, a site for Rosaceae literature, and more analysis tools such as an interactive contig viewer and a comparative map viewer.We plan to incorporate more Rosaceae genomics and genetics data from researchers worldwide as well as data from the ongoing The GDR is initiated to support the genomics and genetics research in Rosaceae, which contains numerous economically important fruit trees and horticultural plants. Currently GDR contains all the genomics data for the Rosaceae model peach, maps and markers of Rosaceae species and all the publicly available Rosaceae ESTs. Our integrated database provides users with easy access and retrieval of the annotated data, and the web tools enable them to further analyze their data. With future plans, including more data acquisition and tool developments, GDR will play an important role in the timely and efficient analysis of the data, the exchange of results and ideas among researchers worldwide, the support of Rosaceae labs worldwide with Bioinformatics tools and the utilization of the data from the model species in the study of other Rosaceae species. The methodology and tools applied to develop GDR should be easily applied to develop other comparative genomic databases for different families..The GDR is publicly available and can be accessed at AFLP: Amplified Fragment Length PolymorphismBAC: Bacterial Artificial ChromosomeCGI: Common Gateway InterfaceEST: Expressed Sequence TagEXP: EXpectation ValueHTML: HyperText Markup LanguageHTTP: HyperText Transfer ProtocolINRA: Institut National de la Recherche AgronomiqueRFLP: Restriction Fragment Length PolymorphismSSR: Simple Sequence RepeatXML: eXtensible Markup LanguageSJ performed web interface design and programming for html pages and JSP search pages, participated in the database construction, carried out database curation, designed the Map Viewer and performed the case study. CJ wrote scripts for database upload and sequence processing, programmed Map Viewer and implemented WebFPC and WebChrom. MS participated in the database construction and wrote scripts for data upload and search pages for Genbank Rosaceae sequences. SF performs database administration. IC developed the application prototype for servlet-database connection and participated in the database construction. AA conceived of the project and participated in its design and coordination. ZD developed the sequence similarity server application. JT participated in the coordination of the project. DM conceived and supervised the project, participated in the database construction, carried out EST analysis, and provided input in the web interface design. All authors read and approved the final manuscript.


PLoS Biology, volume 2, issue 12.In 10.1371/journal.pbio.00203809 beads per g of cells” contains an error. The correct amount of beads is 8 × 108, i.e., ten times less.In Materials and Methods, the sentence “The magnetic beads were added to the extract to a ratio of about 8 ×10Published February 15, 2005