The liver is an important contributor to the human immune system and it plays a pivotal role in the creation of both immunoreactive and tolerogenic conditions. Liver transplantation provides the best chance of survival for both children and adults with liver failure or cancer. With current demand exceeding the number of transplantable livers from donors following brain death, improved knowledge, technical advances and the desire to prevent avoidable deaths has led to the transplantation of organs from living, ABO incompatible (ABOi), cardiac death donors and machine based organ preservation with acceptable results. The liver graft is the most well-tolerated, from an immunological perspective, of all solid organ transplants. Evidence suggests successful cessation of immunosuppression is possible in ~20–40% of liver transplant recipients without immune mediated graft injury, a state known as “operational tolerance.” An immunosuppression free future following liver transplantation is an ambitious but perhaps not unachievable goal. The initial immune response following transplantation is a sterile inflammatory process mediated by the innate system and the mechanisms relate to the preservation-reperfusion process. The severity of this injury is influenced by graft factors and can have significant consequences. There are minimal experimental studies that delineate the differences in the adaptive immune response to the various forms of liver allograft. Apart from ABOi transplants, antibody mediated hyperacute rejection is rare following liver transplant. T-cell mediated rejection is common following liver transplantation and its incidence does not differ between living or deceased donor grafts. Transplantation in the first year of life results in a higher rate of operational tolerance, possibly due to a bias toward Th2 cytokines  during this period. This review further describes the current understanding of the immunological response toward liver allografts and highlight the areas of this topic yet to be fully understood. At present, orthotopic liver transplantation (OLT) is the only effective treatment option for many conditions . UnfortuThe allograft implanted during OLT can be from either a living or deceased donor. Procurement of a deceased individuals organs can occur following brain death (DBD) or cardiac death (DCD) and the organ can be split between two recipients depending on the volume of parenchyma required. Deceased donors are scarce in many countries and implanting organs from different ABO blood groups may be the only option to save a recipient's life, this is known as ABO incompatible (ABOi) liver transplantation. Early reports of ABOi OLT utilizing conventional immunosuppressive regimes and deceased donors yielded significantly inferior results . Howeverliver, transplant, immune response, innate, acquired, cell, antibody, rejection, ischaemia reperfusion, cadaveric, living donor, ABO incompatible, pediatric, pediatric. Any additional publications relevant to this review were then identified by manually searching article reference lists.Relevant existing publications for this narrative review were identified by searching the Pubmed, EMBASE and Medline databases. The search was limited to the English language, but no other filters were utilized. The following terms were utilized (in a variety of combinations); The liver is one of the two organs in human body with a dual blood supply, deriving blood from both arterial and portal venous blood. Therefore, it is exposed to both systemic and enterically derived pathogens . Portal Systemic infection has a significant effect on the liver. Sepsis is known to induce changes in gene and protein expression and this alteration in hepatocellular function is known as the acute phase response. This response is triggered by IL-6 and IL-1 from monocytes and stimulates hepatocytes to release numerous acute phase proteins (APPs) . A numbepos, CD25high, CD127LOW, FoxP3pos), are key components of the immunosuppressive arm of the immune system, suppressing effector cell activity and restoring immune homeostasis. Multiple immunosuppressive mechanisms have been attributed to Tregs such as the secretion of anti-inflammatory cytokines and inhibitor molecules , depletion of crucial growth factors, disruption of effector cell metabolism by promoting the accumulation of adenosine nucleosides, consuming scarce amino acids and also by direct cytotoxic killing of effector cells  is comprised of autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), and primary sclerosing cholangitis (PSC). AIH results from a T cell mediated insult on autoantigens and causes a chronic hepatitis with an interface and lobular hepatitic component, however the portal/interface component usually predominate . A subseor cells . The numor cells . Howeveror cells , 20. Sheor cells . Recruitor cells . The recor cells , 21. Theor cells . The graor cells .A hepatic allograft has immunoprotective benefits. The frequency of renal allograft TCMR is significantly lower in combined liver-kidney recipients in comparison to kidney alone recipients . In addiThe first liver transplant recipient to survive more than 24 h following the procedure was a 19-month-old infant who received a whole liver graft obtained from a 18-month-old brain dead donor . This piA liver obtained from a deceased individual is known as a deceased donor graft and depending on the terminal event, these donors can be considered to have experienced brain death (DBD) or circulatory death (DCD). The universal definition of brain death is “The irreversible loss of the capacity for consciousness, combined with irreversible loss of the capacity to breathe and therefore irreversible cessation of the integrative function of the brainstem” and strict neurological criteria need to be satisfied to make this diagnosis . DBD donLiving donor liver transplantation (LDLT) occurs when a live individual undergoes a partial hepatectomy and donates this portion of their liver. The type of hepatectomy will depend on the volume of parenchyma the recipient requires. The donor can be either biologically related or biologically unrelated. All living donor grafts will be partial grafts, the only exception to this would be when “Domino” transplantation occurs. A “Domino” transplant occurs when recipient A undergoes a total hepatectomy and this explanted liver is transplanted into recipient B, recipient A can survive as they receive a separate cadaveric or living donor graft . This stex-situ in cold storage, thus without perfusion or oxygen delivery. These preservation conditions minimize oxidative phosphorylation and reduce metabolic activity to ~10% of the normal rate, the energy of which is mainly derived by anaerobic metabolism , blood loss and alloantigen recognition. Traditionally the liver grafts are preserved tabolism . In additabolism . Re-estatabolism , 35. Thitabolism . Releasetabolism . Enhancetabolism . Advancetabolism . PRI hastabolism , 39. In tabolism . Figure The human immune system is commonly divided into innate and adaptive components with separate effector cells and activation pathways. However, evidence suggests third division of the immune system referred to as “innate-like” exists and is comprised of both B and T lymphocyte subsets . A charaAcute T-cell mediated rejection (TCMR) is the most common immune mediated complication following liver transplantation . Less frde novo autoimmune hepatitis as a form of plasma cell rich rejection and added criteria for the diagnosis of acute and chronic AMR . DSAs may be against HLA antigens, which are the most readily detected by current assays, or non-HLA antigens such as anti-glutathione S transferase (GSTT-1) and anti-angiotensine 2 receptor (de novo DSA formation experienced acute AMR and the average liver fibrosis score was higher in this subjects with DSAs (Acute AMR causes graft dysfunction due to donor specific antibody (DSA) interaction to antigens on the graft. DSAs may be pre-existing (preformed) or develop post-transplant in response to foreign antigen  cytoprotective  and injury gene  expression associated with DCD and DBD grafts, utilizing a living donor liver as a reference . DirectlP = 0.685) (P = 0.84), the early rejection rate (≤30 days) was identical  . Howeverdentical . A case dentical . PRI is dentical . Despitedentical . Mechanidentical . The res+ CD8 T-cells was significantly higher in the perfusate from DBD grafts   . Coexistormation .The lack of suitable deceased donor livers for transplantation and the associated waitlist mortality has prompted the development of living donor liver transplantation (LDLT). The first successful LDLT was performed in Australia in 1989, a female adult donated her left lobe and it was implanted into her to 17-month-old son who suffered from biliary atresia . Followide novo donor specific antibody formation in LDLT in comparison to cadaveric graft recipients (DSA)  . Avoidants (DSA) . It was P = 0.04)  . De Jong = 0.04) . Gene ex = 0.04) . The par = 0.04) .Transplanting organs across the ABO blood groups has for a long time been associated with poor outcomes due to increased graft loss and worse patient survival . The bloAcute AMR is a feared consequence of ABO-I liver transplantation and can often lead to loss of the graft. Numerous interventions have been attempted to mitigate the risk of AMR and these include; preoperative plasmapheresis, splenectomy, local infusions, mycophenolate mofetil, and rituximab  Several p = 0.038) with non-anastomotic biliary strictures occurring in 12 (8.5%) of the ABO incompatible recipients. Interestingly, all 12 underwent liver biopsy and only one case had histopathological evidence of AMR. No recipient of an ABO compatible graft developed a non-anastomotic biliary stricture  in each group (P = 0.001)  . A large= 0.001) . A recip= 0.001) . It has = 0.001) . This is= 0.001) .P = 0.08) (P ≤ 0.01). Kehar et al. found no difference in rejection rates between pediatric patients that received a graft from a genetically related compared with unrelated donor (P = 0.4) (Post-operative frequency of TREGs and IL-4 are higher in pediatric recipients who receive a LD graft in comparison to a deceased donor graft cadaveric . Favorab = 0.08) . Alonso P = 0.4) .There is variation in both the standard post-operative immunosuppression and rejection treatment regimens utilized by transplant centers around the world. The immunosuppressant drug most commonly used long term is the calcineurin inhibitor, tacrolimus, based on evidence of improved efficacy , 110. ThTolerance inducing therapies are showing promising results in clinical studies. In a landmark pilot study, Todo et al. showed that a single post-operative infusion of TREG cells allowed accelerated withdrawal of immunosuppression at 6 months post LDLT and 70% of these individuals achieved operational tolerance . The patThe field of liver transplantation has advanced significantly since the 1960's. Progress has been made in organ preservation, post-operative care, immunosuppression and optimal utility of different grafts to ensure those in need get the best possible access to this lifesaving procedure. Grafts previously not possible such as ABOi LDLT are now commonplace in many centers with acceptable results. Based on existing literature, a similar immune response is elicited to the majority of grafts following implantation, comprising an initial innate inflammatory response due to preservation-reperfusion mechanisms followed by a predominantly cell mediated response. However, contribution from the innate-like and humoral components of the immune system to PRI and graft rejection are becoming increasingly recognized. Grafts from brain dead donors appear to have a higher inflammatory cell infiltrate and cytokine concentration at retrieval than DCD or living donor grafts, however clinical differences in rejection as a result are not evident. It is likely that both cell and antibody mediated injury results in the morbidity associated with chronic rejection. Acute TCMR is common following OLT but undergoing transplant in the first year of life seems protective. Hyperacute graft rejection of the liver is exceedingly rare. Strong clinical evidence that a DBD, DCD or LDLT graft is associated with a lower rate of TCMR is not apparent. Research into tolerance inducing therapies has shown promising results and the results of larger, phase II trial are eagerly awaited.AH was employed by the University Hospital Birmingham NHS Trust as a clinical research fellow.AH and D-CO-B performed the literature search and constructed the manuscript. DN, VR, SW, and MP reviewed and contributed to the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


SME operators in the informal sector of developing economies have a significant influence on their nation's economies through their involvement in international business relationships. However, the existing deficiency in the literature to show empirical relationships between knowledge transfer, from these SMEs and their international business partners, and innovation performance is a significant gap in the strategic management and international business literature. Therefore, this paper explores the link between knowledge transfer and innovation performance of informal economy SMEs that are involved in international business relationships. The study included a survey of 370 owners-managers and managers of small and medium enterprises in Nigeria's informal electronic market. Using Structural Equation Model (AMOS 22) this study shows that knowledge transfer dimensions, such as R&D and social networking, have varying levels of impact on innovation performance of informal sector SMEs. Knowledge transfer from training showed an inverse and insignificant relationship with innovation performance. The study established implications and recommendations that will be useful for theory and practice. Knowledge transfer; Innovation performance; SMEs; Informal economy; R&D; Social networking; Entrepreneurship; Business policy; Management; Business management; Strategic management. Knowledge transfer is the movement of knowledge through a/some channel(s) from one individual or firm to another . It alsoAccording to According to This research, therefore, raises a question about the capacity of SMEs operating in the informal sector of developing economies  involved in international business relationships, to relate with knowledge transfer from their foreign business associates and to translate same into viable new product innovations. More obvious is the fact that there is a shortage in the existing literature that shows empirical relationships and that explain the processes between knowledge transfer and innovation performance in informal economies, such as Nigeria. Nevertheless, the importance of understanding the relationship between knowledge transfer and innovation performance within these developing and informal economies, like Nigeria, cannot be overemphasised. Nigeria, as a world's fast-emerging economy and the largest economy in Africa . Nigeria22.1Several studies in the field of knowledge management and strategic management have investigated different dimensions of organisation and management of knowledge in various economic and industry contexts . There aKnowledge transfer or exchange involves organisational learning transits between a source to a beneficiary for achieving predetermined outcomes . In a br2.2Innovation is the practice of conceptualising and processing of novel ideas that generate commercial values in the form of new products and services. Within the purview of business practices, innovations have an end goal of servicing customers' expectations in new ways . It is aOrganisations innovate across several levels, including products, services, manufacturing designs, service delivery procedures, and managerial processes or in designing the organisational hierarchy . HoweverEarlier research conducted revealed that innovation positively affects the performance of firms . The dependent variable is new product innovation performance and is the ability of SMEs to mobilise technology and skill to innovate new products that meet customer demands. It adapted from Consequently, items in the research instrument adapted from existing research that closely related to the present study; and the research instrument items were modified to fit into the present study. Details on Knowledge transfer variables such as questions on training, including the exposure of SME operators to continuous development to acquire new skills on the job, were gotten from 4.2The sample size for this study derived from using the 4.3This study tested for reliability using Cronbach's alpha test. It associates each measurement item to every other measurement item and finds the average. Cronbach's alpha tests the reliability of a multi-item scale. When the coefficient is either 0.70 or higher, it is acceptable in a social science research study. The reliability coefficient of items used for the present study is 0.871. Hence, the present study established that the scale items were consistent with the intended purpose of what the research set out to measure. Moreover, for the individual instruments the results showed the following: training (α = 0.852), R&D (α = 0.876), social network (α = 0.576) and innovation performance (α = 0.914). The values justify the reliability of the scale items because most values are greater than the benchmark of 0.7. Confirmatory Factor Analysis (CFA) confirmed the validity of the research items. According to This present study also ensured discriminant validity for the constructs. Discriminant validity reflects the extent to which constructs differ from one another. By the general rule, 5A total of 370 copies of questionnaires administered to respondents were filled correctly and returned by the respondents.5.1This study relied on the goodness of fit criteria for any SEM-AMOS analysis as postulated by previous studies. As depicted in Similarly, investments in social networks will move new product innovation performance of SMEs positively upward by β = 0.16. The relationship between training and innovation performance was negative and minute (β = -0.09). Although this relationship is contrary to expectation, yet it may be indicative of the fact that informal sector SMEs rarely engage in any form of training relationships with their foreign partners. Moreover, the result signifies that training does not make any significant contribution to the SMEs new product innovation performance. The results, also established that informal relationships and knowledge sharing acquired from social interactions, collaborations and strategic alliances built from social networks instead prove to be more effective than any procedural training activities.The present result from this study is significant for improving the performance of Nigeria's SMEs sector. Currently, the SMEs sector accounts for about forty-eight per cent of national GDP and about ninety-six per cent of the total number of businesses in Nigeria, employing eighty-four per cent of the total population . Despite6This study focused on investigating the relationships between knowledge transfer and innovation performance of Small and Medium Enterprises (SMEs) in the Informal Sector of a developing economy, precisely Nigeria. Specifically, this research work raised three hypotheses from the review of the literature. The first hypothesis suggested a possible impact of training on innovation performance of the SMEs. Based on the results from the structural equation modelling analysis, training did not have a significant influence on SMEs' innovation performance. Hence, the emphasis on training as a strategic knowledge transfer tool for SMEs involved in international businesses may not necessarily enhance innovation performance. This result may suggest that SMEs operators in the informal economy of developing nations like Nigeria are more particular about real-life engagement with their businesses and learning through practical meetings with their international business associates that engaging with organised training programmes.Moreover, the fact that most operators in this sector do not possess formal education could also be a resultant factor. The assertion is such that the SMEs operators in this sector have developed business skills through informal apprenticeships, rather than through organised training. Consequently, the result of this study demonstrates a different perspective from studies in more developed economies suggest that training enhances the survival rate of SMEs . The assThe second hypothesis for this study proposed an impact of research and development on innovation performance of SMEs operating in the informal sector. Based on the statistical analysis carried out, the result revealed that R&D positively impacted on new products development. Hence, SMEs in the informal economy of developing nations like Nigeria, which are involved in international businesses can support their innovation performance objectives by adoption R&D as a viable tool. The third hypothesis was to identify the impact of social networking on innovation performance. Based on the structural equation modelling carried out, the result revealed that social networking impacts positively and significantly on the innovation performance of SMEs operating in the informal economy. Consequently, knowledge transfer that occurs during social networking that occurs between the SMEs and their international business associates help these SMEs to achieve technology breakthrough and new market breakthrough for their businesses. In line with this finding, existing research suggests that firms use social networking to develop connections through online communication, thereby empowering them to improve areas of concern, especially about technology adoption and enhancing their market competitiveness .7This study has adequately shown that knowledge transfer significantly impacts on the innovation performance of small and medium enterprises operating in the informal economy of Nigeria. Specifically, this study has made an original contribution to knowledge management and international business literature by examining SMEs in the developing informal economies that are involved in international business relationships. Based on the findings from this study, it this study asserts that SMEs operators should intensify on R&D as a way of keeping themselves updated on the latest developments in the market and the changing customer trends, to innovate according to identified patterns. Also, SMEs operators should endeavour to establish social networks both among themselves, to facilitate learning and current innovations, and with their international associates. The benefits derivable from such global social networks would include innovating with price innovation mechanisms, for example acquiring products at lower lost. Another important implication from this study is that policymakers and investors should focus attention on encouraging and funding R&D and social networking activities to improve innovations among the SMEs in the informal economy.Theoretically, this study has advanced reasoning in the knowledge management and international business literature about the impact of knowledge transfer and new product innovation performance of informal economy SMEs that are involved in international business relationships. The findings from this study establish a platform for further studies to be replicated in new developing economies and using a cross country analysis. The conclusions of this study apply to a larger group of SME operators in developing economies across the globe and such economies with similar characteristics with a growing informal economy. Hitherto, the literature consists of explanations relating to knowledge transfer among SMEs in international business from a developed economy perspective. The understanding of the characteristics and potentials of informal sector SMEs in international business relationships from a developing economy perspective has been an unfilled gap in the literature. Therefore, this study establishes empirical evidence about the relationship between knowledge transfer and new product innovation performance of informal sector SMEs in Nigeria.8In this study, the focus was on investigating the impact of knowledge transfer on innovation performance of small and medium enterprises in an informal economy. Following the findings from the structural equation modelling, the study concludes that knowledge transfer practices, especially R&D and social networking, have a significant impact on the innovation performance of informal economy SMEs that are involved in international business relationships. This study has advanced insights on the importance of stakeholders in the SMEs sector of informal economies to advance policies, funding and practices of R&D and social networking to improve SMEs innovative performance. Nonetheless, this study was limited to a single informal economy. Despite that Nigeria constitutes a substantial informal market in the global informal economy. Nevertheless, findings from future studies that will adopt a cross-country analysis may supply more robust insights.A. S. Ibidunni: Conceived and designed the experiments; Performed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data; Wrote the paper.A. Kolawole: Conceived and designed the experiments; Contributed reagents, materials, analysis tools or data; Wrote the paper.M.A. Olokundun and M. Ogbari: Contributed reagents, materials, analysis tools or data; Wrote the paper.10.13039/501100012497Covenant University Centre for Research, Innovation and Discovery.This work was supported by The authors declare no conflict of interest.No additional information is available for this paper.


Triticum aestivum) provides 20% of the calories consumed daily across the globe. The nutrients in the wheat grain come primarily from the starch composed of amylose and amylopectin. Resistant starch content, which is known to have significant human health benefits, can be increased by modifying starch synthesis pathways. Starch synthase enzyme SSIIa, also known as starch granule protein isoform-1 (SGP-1), is integral to the biosynthesis of the branched and readily digestible glucose polymer amylopectin. The goal of this work was to develop a triple null mutant genotype for SSIIa locus in the elite hard red winter wheat variety ‘Jagger’ and evaluate the effect of the knock-out mutations on resistant starch content in grains with respect to wild type.Lack of nutritionally appropriate foods is one of the leading causes of obesity in the US and worldwide. Wheat (SSIIa in the three genomes of wheat variety ‘Jagger’ were identified using TILLING. Subsequently, these loss-of function mutations on A, B, and D genomes were combined by crossing to generate a triple knockout mutant genotype Jag-ssiia-∆ABD. The Jag-ssiia-∆ABD had an amylose content of 35.70% compared to 31.15% in Jagger, leading to ~ 118% increase in resistant starch in the Jag-ssiia-∆ABD genotype of Jagger wheat. The single individual genome mutations also had various effects on starch composition.Knock-out mutations in ssiia-∆ABD mutant showed a significant increase in RS without the shriveled grain phenotype seen in other ssiia knockouts in elite wheat cultivars. Moreover, this study shows the potential for developing nutritionally improved foods in a non-GM approach. Since all the mutants have been developed in an elite wheat cultivar, their adoption in production and supply will be feasible in future.Our full null Jag-The online version contains supplementary material available at 10.1186/s12870-020-02822-5. Around 13% of the people in the world are obese and according to available data, obesity is one of the largest risk factors for human deaths globally . In 2017The calories and nutrition that are associated with wheat products come primarily from starch, which serves as the energy and carbon reservoir for the embryo. Roughly 70–90% of the dry weight of the endosperm is composed of starch , 6. StarStudies on RS have shown incredible benefits to human health ranging from maintaining healthy bowel function to helping moderate glycemic indices, as well as the possibility to prevent colon cancer –14. RS iTriticum durum) (genome AABB) and bread wheat (genome AABBDD), have shown an increase in amylose [Starch synthesis within the endosperm of wheat is accomplished by several enzymes, with differing roles in the elongation and branching of glucose polymers. Granule-bound starch synthase (GBSS) isoforms are the sole enzymes responsible for synthesis of amylose in starch formation . Amylose amylose , 28, 29. amylose , 25, 30. amylose , 32. Stu amylose , 33–35.In this study, an EMS (ethyl methanesulfonate) mutagenized TILLING  population of hard red winter bread wheat variety ‘Jagger’ was used to study the impact of knock-out mutations of starch synthase genes on the RS and amylose content . Jagger SSIIa gene homoeologs were identified in “A”, “B”, and “D” sub genomes and combined by crossing to develop a full null ssiia mutant. With increased RS content in these individuals, we show the ability to develop increased nutritional value in an important cereal crop without the use of transgenics.With the use of our Jagger TILLING population, knockout mutants of Two predictive knock-out mutants each were found for B and D genomes, whereas only one was found for A genome  genotypes. Interestingly, Jag-ssiia-∆B seeds were widest among all the mutant combinations studied, although they did not show any significant difference in grain width in comparison with WT. Between genotypes, Jag-ssiia-∆B showed a significant increase (P < 0.0001) in grain width in comparison with Jag-ssiia-∆A, ssiia ∆ D, and Jag-ssiia-∆ABD  difference between the genotypes . Similarly, all other genotypes showed a significant decrease (P < 0.00001) in TGW in comparison with WT as well. A 21.29% decrease in TGW on average was observed between WT and Jag-ssiia-∆ABD. Surprisingly, Jag-ssiia-∆B had a slightly, but significantly (P < 0.01) higher TGW than Jag-ssiia-∆A, ssiia ∆ D, and Jag-ssiia-∆ABD, which is consistent with grain width data.In thousand grain weight (TGW), a significant decrease was observed between Jag-ssiia-∆ABD in comparison with WT (P < 0.01), with WT having an amylose content of 31.15% on average, and 35.70% on average in Jag-ssiia-∆ABD. Interestingly, Jag-ssiia-∆D had a significant decrease in amylose content in comparison to WT (P < 0.001). Similarly, Jag-ssiia-∆ABD had a significant decrease in total starch (TS) (P < 0.0001) in comparison with WT. Additionally, Jag-ssiia-∆D showed no significant difference in total starch in comparison to WT. However a significant increase (P < 0.0001) in TS was observed between Jag-ssiia-∆D and Jag-ssiia-∆ABD , a total of a 118.81% increase in RS. Significant increases (P < 0.001) of RS percent were also observed between Jag-ssiia-∆ABD and every other genotype . Between genotypes, Jag-ssiia-∆ABD was shown to have a significant increase (P < 0.01) in RS content in comparison with the other mutant genotypes  was observed between percentage RS and amylose content. However, a moderate negative correlation was observed between percentage RS and TS (r = − 0.68). RS content (g/100 g) followed a similar pattern as percentage of resistant starch, but to a lesser degree. A strong positive correlation (r = 0.76) was observed between RS content and amylose content, and a moderate negative correlation (r = − 0.55) was observed between RS content and TS. A moderate negative correlation (r = − 0.63) was observed between amylose content and TS Fig. A strong positive correlation (r = − 0.63), as well as between percentage RS and TGW (r = − 0.58). A strong negative correlation was observed between percentage RS and grain length (r = − 0.86). TS and grain length showed a strong positive correlation (r = 0.82). Similar correlations were observed between RS content (g/100 g) and grain width (r = − 0.64), grain length (r = − 0.81), and TGW (r = − 0.47). Interestingly, amylose content showed no significant correlation (P > 0.01) with grain width. A weak positive correlation was found between TS and grain width (r = 0.46), and a strong positive correlation was found between total starch and TGW (r = 0.80).Moderately negative correlations were observed between percentage RS and grain width , cv Turkey, which naturally lacked the ssiia gene in the D genome, was used to integrate a null D genome ssiia gene into a full null individual [SSIIa genes in all three genomes and were used as controls [SSIIa homoeoalleles in Jagger or background mutations in the D genome mutant could be the underlying factors for the reverse effect on amylose content and TS of the ssiia D genome knock-out mutant. Further studies will be needed to identify the exact reason for this unexpected trend with the D genome copy of ssiia.In this study, we have developed a full null id wheat –32, 34.  genomes . The Jagdividual . No signcontrols . Non-balssiia null mutant durum wheat, which is generally used for pasta flour, studies have shown a decrease in cooking time, increased firmness, and a resistant to overcooking in pasta. This, in addition to a higher protein and fat content are indicators of beneficial properties of flour derived from ssiia mutant wheat [ssiia null mutants. Field-based experiments will be done in future once the background has been cleared of additional mutations by back-crossing to see if a detectable reduction in yield is observable due to exclusively the ssiia mutations. Additionally, some cooking properties have been shown to be negatively affected by high amylose content. Studies focusing on SBE genes in in durum and bread wheat have shown significant increases of 55.13 and 108.77% on average respectively in amylose content [P = − 0.53) between amylose content and milling flour yield [-ssiia-∆ABD mutant can be used as a specialty wheat for targeting weight management and other health benefits in humans, however additional studies on non-starch polysaccharide (NSP) and sugar content may give further insight on the nutritional benefits of these grains.In studies of nt wheat . In addint wheat . However content , 42, 45. content , 31, 33. content . In an aur yield . Howeverssiia-∆ABD mutant was 118.81% higher than WT, which is significantly lower than studies on SBE gene knockouts. On average, SBE null mutants had an RS increase of roughly 652.95% in durum wheat, and 1132.39% in bread wheat [ssiia null individuals were developed by Botticella et al. (2016), and showed an increase in RS of 645% on average [ssiia mutants were developed by crossing two different varieties that had null ssiia genes in the A and B sub genomes and were highly shrivelled.The resistant starch content in our Jag-ad wheat , 45, 47.ad wheat , 25. In  average . It is iSSIIa mutant in a single winter wheat background. It further exemplifies the power behind TILLING as a genetic tool. The benefits with working with a hexaploid wheat TILLING population are that a significantly smaller population of mutant individuals  are required to get a knockout of any gene of interest. Tetraploid and diploid wheat species require significantly larger populations to have the same effect, ~ 3000 and ~ 5500 individuals, respectively [ssiia null individuals harbor several background mutations, but these can be significantly reduced through two to three rounds of back-crossing with the use of marker assisted breeding.This study marks the first development of a full null ectively , 40. We ectively  and the ssiia-∆ABD of an important gene in the starch synthesis pathway was developed in this study. The starch composition is similar to that of full null mutants in tetraploid backgrounds created through traditional breeding. In addition, seeds of Jag-ssiia-∆ABD genotype, though having a significant loss in TGW, still do not show a shriveled phenotype unlike previous hexaploid and durum SSIIa mutants. Subsequent experiments involving starch granule and overall grain morphology may give insight on why this is the case. The Jagger ssiia mutants are all in a single elite genetic background, making the phenotype less affected by compounding genes from different genetic backgrounds. However, it is important to note that the predictive combined mutants have not been back-crossed so far, leading to several background mutations not present in parental WT Jagger. The Jagger ssiia null mutants showed a significant increase in RS, a nutritionally important trait in cereals that has shown several health benefits in humans. Though the increase of RS was relatively small (from ~ 1% to ~ 2%) this still shows the functionality of these amylopectin synthesis genes, and how their modification can affect, and ultimately improve these nutritionally important traits. As with any modification in amylopectin biosynthesis pathway, these ssiia null mutants had less total starch, and grain width was reduced. Interestingly, the Jag-ssiia-∆B and Jag-ssiia-∆D mutants showed phenotypic variances that pique an interest and may give insight on the roles of the different homoeologs of SSIIa in wheat. Grains of Jag-ssiia-∆B had no significant difference in their width, though TGW was still significantly lower than wild type yet significantly higher than the full null mutant. Mutant Jag-ssiia-∆D had a seemingly reverse phenotype in amylose content and total starch content, with amylose percentage being significantly lower than the ssiia null mutant as well as WT, in addition, and as is the trend, total starch percentage was significantly higher than the ssiia null. This study shows the possibility of developing non-GM, nutritionally superior cultivars in an efficient manner. Established, elite cultivars, such as Jagger, are already widely accepted and used by growers, and modifications of single gene for increasing its nutritional value will lead to its easy adoption in production and supply chain as a specialty wheat. This study shows the impact of the modification of one gene in the starch synthesis pathway, and the possibilities of the health benefits that coincide with these modifications.A triple knockout mutant Jag-1 plants were obtained from the treated (M0) and planted in a 1:1 vermiculite/soil mixture. Vernalization was done at the two-leaf stage for 6 weeks at 4 °C. M1 Plants were allowed to self, resulting in M2 seeds. Out of the 1310 M2 seeds planted, 1296 M2 plants germinated, each with a unique set of SNPs. M2 plants were used for tissue collection, and high throughput DNA extraction was done using Quiagen Biosprint 96 robot with a Biosprint 96 Plant DNA extraction kit (Qiagen) according to the manufacturer’s instruction. DNA quantification was done using a Nanodrop . Dilutions of 25 ng/μL were made in 96-well blocks, and 4X DNA pools were developed by combining 200 μL of normalized DNA from four 96-well blocks, retaining column and row identity. All the plants were grown in greenhouse unto maturity for 16:8 h of day:night length at 22 °C: 18 °C day and night temperatures. The plant research conducted in the work is compliant with institutional, national and international guidelines.Development and characterization of the Jagger TILLING population used in the work has been described in detail in Rawat et al. (2019) . The plaSSIIa) gene were obtained from Shimbata et al. (2005) [SSIIa-A, SSIIa-B, SSIIa-D) of the SSIIa gene keeping the reverse primer common, designed by Shimbata et al. (2005)  Pools that contained a mutation were then deconvoluted by performing a similar procedure with each individual within the target pool [2 DNA and 2) with equal proportions WT Jagger DNA and M2 DNA. This allowed for the determination of zygosity of the mutation within the individual. Homozygous mutations appeared only in sample 2 which had WT + mutant DNA, whereas heterozygous mutations appeared as a digested band in both the reactions. Sanger sequencing was done to determine the mutation in the confirmed mutant individuals  . Clustal5) Table . Figure  . Clustaget pool . Two rea . ClustaCel-1 based zygosity determination was done as described above. Individuals with homozygous mutations were selected and crosses were made as A x B, and B x D to generate F1 hybrids with mutations in A-B and B-D homoeoalleles. F1 seeds were harvested and planted. Mutations were confirmed by Sanger sequencing the A-B and B-D homoeoalleles of SSIIa in the F1 plants to rule out self-pollination. A double cross was made between these F1 plants to combine all three mutations, retaining the B genome mutation in homozygous condition. Sixty-nine seeds were obtained from the double cross, which were planted again. Zygosity determination was done, and 4 plants were found to have B genome mutation in homozygous condition, and A and D genome mutation in heterozygous condition. These plants were selected and allowed to self. A set of 96 seeds were planted from the bulked selfed seeds, DNA was extracted and zygosity was determined using Cel-1 assays. Six plants were found to contain all A, B, and D mutations in homozygous state. Mutations were confirmed by Sanger sequencing in all three genomes. These six plants were allowed to self and seeds were bulked for further analysis of starch composition. Homozygous A, B, and D genome mutants in addition to Wild type Jagger plants were used to compare the parameters of starch composition and grain size comparisons.Plants with knockout mutations in each of the A, B, and D genomes were grown, DNA was extracted, and n = 25) was done using free software ImageJ. After determining homogeneity of variances using Levene Test as well as testing for normality using the Shapiro-Wilk Test, statistical analysis of the results was done using one-way ANOVA, followed by Tukey T-test using R. Correlation analysis was done using the corrplot package in R.One gram of whole dry seeds was manually ground in a mortar pestle to a fine flour for each genotype. Amylose content was determined using Megazyme International’s Amylose/Amylopectin Assay Kit (K-AMYL) as per the manufacturer’s protocol. Six biological reps with two technical reps were preformed with the amylose/amylopectin assay kit. Resistant starch as well as total starch content was determined using Megazyme International’s Resistant Starch Assay Kit (K-RSTAR) as per the manufacturer’s protocol. Three biological reps with two technical reps were preformed with the RS assay kit. Measurements of grain size (The plant research conducted in the work is compliant with institutional, national and international guidelines.Additional file 1: Supplementary Table 1. DNA amplicon sequences of wild type Jagger and truncation mutants for SSIIa gene A, B and D genome copies. The red highlighted bases in the mutant sequences in the table indicate the variation of gene sequence in them. Supplementary Table 2. Partial protein sequences of wild type Jagger and truncation mutants deduced from the amplicons for SSIIa A, B and D genome copies.


The specimens were rapidly cooled from high temperature inducing a diffusionless cubic-to-metastable tetragonal (t’) phase transformation. The changes in *a*b*L color coordinates, translucency parameter (TP), and total transmittance (T%) were measured. Three-point bending strength, Vickers hardness, and indentation fracture toughness tests were performed. Quantitative phase analyses were carried out by X-ray diffraction with Rietveld refinement. Scanning electron microscopy (SEM) images were obtained. With increasing Y2O3 contents, TP and T% values increased while strength and toughness decreased. The Rietveld analysis showed that the amount of t’-phase increased after rapid-cooling and annealed 5Y-partially stabilized zirconia (PSZ) contained the highest amount of t’-phase (64.4 wt%). Rapid-cooling improved translucency but the translucency of annealed 5Y-PSZ did not approach that of lithium disilicate glass-ceramic. Rapid-cooling decreased flexural strength significantly, being 306.1 ± 61.8 MPa for annealed 5Y-PSZ. SEM revealed that grains tended to get larger after rapid-cooling. A rapid-cooling treatment can produce t’-phase which can contribute to an increase in translucency but has a negative effect on the mechanical properties of zirconia.Many attempts have been made to improve the translucency of zirconia in dentistry. The purpose of this study was to evaluate the effect of a rapid-cooling heat treatment on the optical and mechanical properties of dental monolithic zirconia. Zirconia containing 3, 4, and 5 mol% Y The. The7]. *L = 94.26, *a = −0.24, *b = 0.95), a black glass-ceramic tile , and an A2 glass-ceramic tile .For sintered/polished thin plate specimens, spectral reflectance data in the wavelength range of 360–750 nm at 10-nm intervals were acquired with a spectrophotometer  against a white polytetrafluoroethylene background  and diffuse/8-degree geometry were calculated from the reflectance data. A 6-mm diameter aperture and a 6-mm diameter measurement area were used. The measurements were conducted by using glycerin as an optical coupling medium in-between the specimen and the background . The tra31 and diThe total transmittance (T%) was calculated using the following formula :(2)T%= method on the thick plate specimen using a Vickers microhardness tester  with a load of 10 kg for 10 s. Five indentations were used to determine the average hardness. Flexural strengths of the bar specimens were measured by using a 3-point bending test with a 30-mm span according to International Organization for Standardization standard 6872:2015 . The speTo identify the crystalline phase of zirconia specimens, one specimen from each zirconia group (sintered/polished or rapidly-cooled) was randomly selected. XRD  was conducted by using Cu-Kα radiation (λ = 1.54056Å) at 40 kV and 40 mA. XRD profiles were collected between 25 and 80° (2θ) at a scan speed of 1°/min in a continuous-scanning mode with a step size of 0.01°.2/nmc, monoclinic zirconia unit cell with space group P21/c, cubic zirconia unit cell with space group FmThe relative phase contents of monoclinic (m), tetragonal (t), and cubic zirconia (c) were determined by Rietveld refinements with Topas academic software . The phase structures were refined as: tetragonal zirconia unit cell with space group P4−5 Pa) with an acceleration voltage of 15.0 kV and an emission current of 64.2 µA.The microstructures of the thermally-etched (zirconia) or acid-etched (lithium disilicate) specimens were observed using a scanning electron microscope . For zirconia, thermal etching was carried out at 1350 °C for 30 min with fast heating rate (20 °C/min) to prevent grain growth. After coating with a thin layer of platinum , secondary electron SEM images were acquired in vacuum . The Kolmogorov-Smirnov test was done to determine the normality assumption. A two-way ANOVA was performed to assess the interrelationship of two independent variables (yttria content and rapid-cooling heat treatment) on the optical or mechanical properties of zirconia. The significance level was set at 0.05.*L, *a, and *b against an A2 background, TP, and T% at 550 nm of each group are reported in *a; F  = 170.412, p < 0.001, partial η2 = 0.662, observed power = 1.000 and *b; F  = 58.798, p < 0.001, partial η2 = 0.403, observed power = 1.000. Simple main effects analyses showed that there were significant differences in *a values between sintered/polished and rapid-cooling groups for 3Y- and 4Y-PSZ (p < 0.001) and there were significant differences in *a values among 3Y-, 4Y-, and 5Y- groups for sintered/polished (p < 0.001) and rapid cooling groups (0.001 < p < 0.006). Simple main effects analyses showed that there were no significant differences in *b values among 3Y-, 4Y-, and 5Y- groups for rapid cooling groups (0.157 < p < 0.798). There were no significant interactions between the effect of yttria content and a rapid-cooling heat treatment on *L; F  = 0.094, p = 0.911, partial η2 = 0.001, observed power = 0.064, TP; F  = 0.164, p = 0.849, partial η2 = 0.002, observed power = 0.075, and T%; F  = 2.288, p = 0.104, partial η2 = 0.026, observed power = 0.460. According to Tukey post hoc test, *L value decreased, TP value increased, and T% increased as the yttria contents increased for sintered/polished. The additional rapid cooling treatment for each zirconia group improved translucency, as indicated in Means of CIE pThe mechanical properties of each ceramic group are reported in c/a ratio  of the t’-phase decreased, approaching unity. Rapid-cooling of the sample caused an increase in the t’-phase contents and R/5Y-PSZ contained the highest amount of t’-phase (64.4 wt%). The results are summarized in The quantitative phase analysis by using the Rietveld method indicated that t’-phases were present in all samples. 3Y-TZP contained 79.2 wt% t-phase and 20.8 wt% t’-phase, and the t’-phase increased to 59.1 wt% for 5Y-PSZ. As the yttria contents increased, the SEM images of each ceramic group were shown in 2O3. Therefore, the null hypothesis was rejected.In order to further improve the translucency of dental monolithic zirconia, the author fabricated t’-phase-containing translucent zirconia. According to the results of this study, a rapid-cooling protocol found to be detrimental to the mechanical properties even though it improved the translucency of dental monolithic zirconia containing 3–5 mol% Y2O3-ZrO2 samples caused the formation of new domains with different texture (t’-phase). The t’-phase was induced by the displacement of oxygen ions in the lattice parameter through a diffusionless mechanism. The presence of t’-phase has been verified by its characteristic platelike domain structure with antiphase boundaries [2 and the weight fractions of each phase were quantified using the Rietveld refinement method. Zhang and Lawn [2O3-ZrO2 samples had two types of tetragonal phases (t + t’) and no monoclinic or cubic phases were identified. With increasing yttria contents (from 3–5 mol%), the proportion of t’-phase increased at the expense of t-phase. The t’-phase is morphologically different from t-phase even though crystallographically identical. In addition, c/a ratio  of t’-phase approached unity with increasing yttria contents. Due to the close proximity of its crystal parameter to the isotropic structure, light scattering from birefringence can be reduced resulting in enhanced translucency of dental zirconia. In this study, annealing at 1550 °C allowed the material to reach its equilibrium state and upon rapid cooling, no t → m transformation occurred. From the c + t two-phase region, c-phase went through a diffusionless c → t’ transition and the t-phase stayed up to room temperature. By a rapid-cooling heat treatment protocol, the t’-ZrO2 was formed for all zirconia groups and thus, the enhanced translucency was obtained.Jue et al.  reportedundaries . Due to undaries , it was and Lawn  reportedSince cubic zirconia do not undergo stress-induced transformation , it adveBased on the results of this study, increased yttria contents reduced the lightness color value of zirconia. In addition, a rapid-cooling heat treatment also affected color values of zirconia. The lightness increased, while the chroma decreased. The translucency was improved by a rapid-cooling heat treatment although the value did not reach that of lithium disilicate. Unlike the t-phase, the stable t’-phase does not suffer from long-term thermal aging which causes a degradation of the mechanical properties .2O3-ZrO2. However, according to the results of this study, a rapid temperature drop in the cooling process weakened the mechanical properties. Therefore, more effective cooling protocols to yield t’-phase without sacrificing high toughness should be investigated. After heating at the temperature inside the two-phase field (following Scott [The application of the t’-phase would be one of the strategies to increase the translucency of Yng Scott ), coolinThe present study has some limitations. Indentation Fracture (IF) method was used to test fracture toughness. A recent study  reported2O3 contained t’-phase instead of c-phase, as observed by the Rietveld analysis. Particularly, the author tried to produce t’-phase through a rapid cooling protocol according to the temperature-dependent phase transition mechanism of zirconia. Based on the results obtained, the formation of t’-phase increased the translucency of zirconia and t’-phase-containing translucent monolithic zirconia could be fabricated not with an addition of dopant element but with the change of phase composition through a specific cooling protocol.Although there have been much research on the fabrication of high-translucent monolithic zirconia with an addition of dopant element, there have been no research on the production of t’-phase-containing translucent zirconia. Due to the high overlapping diffraction pattern with c-phase reflection, t’-phase can be easily missed. In this study, it was found that sintered/polished dental monolithic zirconia with 3–5 mol% YIn summary, the author fabricated t’-phase-containing translucent monolithic zirconia by altering phase compositions using a rapid-cooling protocol. The t’-phase can improve the translucency, while its mechanical properties were sacrificed. The optimal protocol to improve the translucency without deteriorating its mechanical characteristics of dental monolithic zirconia will be determined in the future.


Two different starch samples were used. The first starch (CS1), despite extensive heating and shearing contained “ghost” granules, while the second starch (CS2) had no visible ghost granules after the same gelatinization process as CS1. Apparent viscosity measurements demonstrated that κC + CS1 mixtures were shear thinning liquids, with viscosity values being lower than the corresponding weight average of the values of the individual equilibrium phases at shear rates < 50 s−1. Tribological results revealed that κC ≥ 0.5 wt% was required to observe any decrease in friction coefficients in the mixed lubrication regime. Starch (CS1) showed an unusual behavior at ≥ 5 wt%, where the friction coefficient decreased not only in the mixed regime but also in the boundary regime, probably due to the presence of the “ghost” granules, as the latter became entrained in the contact region. The CS1 + κC mixtures showed significantly lower friction coefficients than that of pure CS1 and κC in the mixed regime. However, the CS2 + κC mixture  showed similar behavior to pure κC in the mixed regime, while lower friction coefficients than that of the pure CS2 and κC in the boundary regime. These findings illustrate new opportunities for designing biopolymer mixtures with tunable lubrication performance, via optimizing the concentrations of the individual biopolymers and the gelatinization state of the starch.In this study, we investigated the rheological and tribological properties of biopolymer mixtures of gelatinized corn starches (0.5 – 10.0 wt%) and  Torres et al.  have been investigated , highlighting their ability to entrain and prevent direct PDMS–PDMS asperity contact, irrespective of the degree of surface roughness  of these intact starch granules on the tribological properties of the biopolymer mixtures. Ghost granules, or ghost remnants, are derived from the external layers of the granules of various starches, depending on the ratio of amylose /amylopectin present in the starch. They are shown to exhibit elastic/plastic properties, therefore, they are likely to affect oral processing and even lubrication  and κC and the findings bring new knowledge for the design of biopolymer mixtures with tailored lubrication performance.To our knowledge, this is the first study that reports the frictional properties of biopolymer mixtures of CS and 22.1κ‐carrageenan (κC), product code 22048 (CAS number 11114‐20‐8), corn starch (CS1), product code S9679 (CAS number 9005‐25‐8) and the second corn starch sample (CS2) 10120 (CAS number 9037‐22‐3), both derived from maize, were all purchased from Sigma‐Aldrich, Dorset, United Kingdom. Although both CS1 and CS2 contain mostly amylopectin, CS1 contained residual ghost granules after heating which was not there in CS2. Polysaccharide solutions were prepared in 20 mM phosphate buffer at pH 7.0. Smooth polydimethylsiloxane  tribo‐couples, that is, ball (∅ 47 mm) and disc , with the surface roughness of 50 nm, were purchased from PCS Instruments, London, United Kingdom. Rhodamine B (product code R‐6626) was purchased from Sigma Aldrich. Water purified by a Milli‐Q apparatus , with a resistivity not less than 18.2 MΩ.cm, was used for the preparation of the buffer and any other solutions.2.2. κ‐carrageenan (κC) was similarly dispersed in phosphate buffer at pH 7.0 (0.05 – 1.0 wt%) for at least 24 h at room temperature and then heated at 90°C for 20 min. To prepare the biopolymer mixtures, both solutions were prepared separately as above before mixing. Equal volumes of starch solution and κC solution of different concentrations were blended and homogenized at 21,000 rpm for 30 min using an Ultra Turrax T25 homogenizer  at room temperature (25°C).Gelatinized waxy corn starch (CS1 or CS2) (0.5 – 5 wt%) was prepared by dispersing the starch powder in phosphate buffer at pH 7.0, followed by heating in a water bath at 90°C for 20 min with constant shearing using a magnetic stirrer, to gelatinize the starch2.3−1 at 37°C. The biopolymer mixtures did not phase separate within the first 2 h of preparation but started to separate after 7 days of storage at ambient conditions  at shear rates ranging from 0.1 to 1,000 sions see  and all 2.4W) of 2 N and a slide‐to‐roll ratio (SRR) of 50% were set for all Stribeck measurements. The sliding speeds were varied from 1 to 0.001 m s‐1. The coefficient of friction was measured for all samples as a function of entrainment speed. The entrainment speed U is defined as in Equation UB is the rolling speed of the ball and UD is the sliding speed of the disc. The temperature in the tribological experiment was set at 37 ± 1°C, matching that of the rheological measurements. In addition, the load‐bearing ability of the biopolymers was also measured at loads (W) ranging from 1 to 5 N, at U = 0.005 m s‐1. The friction coefficients are reported as the mean and SD of 3 measurements carried out on at least triplicate samples prepared on different days.Tribological measurements on the pure biopolymers and their mixtures samples were performed using a Mini Traction Machine 2  with hydrophobic polydimethylsiloxane (PDMS) ball and disc as tribopairs. All the tribological experiments were carried out within 2 h of preparation of the biopolymer mixtures where the mixtures were in a single phase . A norma2.5Optical microscopy  was used to observe the microstructure of the heated starch samples to identify any residual granule structures. Samples were diluted with buffer (1:10 wt/wt). A confocal laser scanning microscope  was also used for microstructural characterization of some samples, after mixing with 0.5 wt% Rhodamine Blue (RB), excited at 514 nm, to fluorescently label the starch. Samples were excited with a He/Ne  laser source. A 20× objective with numerical aperture 0.5 was used to obtain all images, at 1,024 × 1,024 pixel resolution.2.6SDs of three measurements on triplicate samples (n = 3 × 3). The statistical analyses were conducted for the rheological data at 50 s−1 simulating oral processing shear and tribological data at boundary regime (0.005 m/s) and mixed regimes  using one‐way analysis of variance and multiple comparison test using SPSS software  and the significant difference between samples were considered when p < .05 using Tukey's test. Statistical results can be observed in All experimental results were reported as mean and 33.1μ) as a function of entrainment speed (U) of κC (0.05 – 1.0 wt%) and CS1 (0.5 – 5.0 wt%), respectively. The phosphate buffer on its own   but no significant decline was observed when the concentration increased from 0.5 wt% to 1.0 wt% (p > .05) . It is noteworthy that κC solutions ≥ 0.5 wt% also demonstrated a characteristic hydrodynamic lubrication regime behavior, where the μ values started to increase at the highest U.The s Figure  showed is et al.  have alrU = 0.1 m/s), but slightly lower μ values in the mixed lubrication regime as compared to that of the buffer. Interestingly, μ was more or less the same for all CS1 concentrations between 0.5 and 2.0 wt% irrespective of the regimes (p > .05)  (see U approached the maximum applied (U = 1 m s‐1). Particularly CS1 at 5.0 wt% did not show any visible boundary regime. This behavior might be due to the presence of higher concentrations of “ghost” granules at the higher CS1 concentration, which potentially might have granule‐granule inter‐molecular friction, flattened and providing sufficient hydrodynamic lift to separate the PDMS surfaces leading to the immediate onset of the mixed lubrication regime even at low speeds (U ≤ 0.005 m s‐1).Shifting our focus to starch, 0.5 – 2.0 wt% CS1 Figure  showed sC Figure , the lat1 Figure  were sigThe presence of ghost granules in the CS1 sample was evidenced via optical and confocal laser scanning microscopy, as shown in Figure Liu, Stieger, Van Der Linden, and Van De Velde  have repκC and CS1 solutions, respectively. As one might expect, higher viscosity values occurred at higher concentrations of the biopolymers  and the viscosity was significantly higher than those of 0.5 – 2.5 wt% CS1 at the typical oral processing shear rate of 50 s−1  of κC and CS1 as a function of W at low U (0.005 m s‐1) is shown in Figure μ values of κC and CS1 as a function of U at different W in μ for an adsorbed polysaccharide scales with W as W23. This dependency originates from Equation A = the contact area for a circular contact =π3WR*4E*23; τi is the interfacial shear stress between the surfaces. The F versus W data in Figure The load‐bearing capacity of the lubricating film is an important parameter to help understand the lubrication performance of biopolymers. The friction force . Thus CS1 lowers the interfacial shear stress between the surfaces even more than κC suggesting that CS1 is more likely to remain bound to the PDMS surfaces. This further explains the larger decrease in friction coefficient with CS1 and the almost non‐existent boundary lubrication regime illustrated in Figure The dashed lines in Figure 3.3κC mixtures, irrespective of the total biopolymer concentration (1.65 or 2.75 wt%) have non‐Newtonian behavior and the viscosity values at orally relevant shear rate of 50 s−1 were significantly different in comparison to either CS1 or κC (see κC and CS1) were similar to that of the starch alone  at corresponding shear rates, except in case of the 1.65 wt% mixtures at ≥ ca. 50 s−1  except for the 2.75 wt% mixture (p < .05)   Figure aii,bii. ) Figure .μ values versus U of a mixture of 2.5 wt% CS2 + 0.25 wt% κC, that is, the starch which contained no starch granules. Similar to the mixtures containing CS1     still contained ghost granules while the other (CS2) did not. In tribological measurements, κC showed good lubrication performance by efficiently reducing the friction coefficient in the mixed lubrication regime, especially when the concentration was ≥ 0.5 wt% and also showed hydrodynamic behavior at higher entrainment speeds (U). On the other hand, the CS1 at ≥ 5 wt% immediately showed onset of a mixed lubrication regime at U < 0.01 m s‐1 without any observed boundary regime. This was attributed to the presence of “ghost” starch granules that flattened in the confinement and enabled accelerated onset of the mixed lubrication regime, which also provided better load‐bearing ability than κC. Mixtures of CS1 + κC (at 1:10 wt/wt ratio) showed that the viscosity values were lower than the weight average of the individual equilibrium phases (CS1 + κC) and the μ values of the mixtures containing the ghost starch granules (from CS1) were much lower in the mixed lubrication regime. The mixture CS2 + κC, lacking ghost starch granules, did not offer such lubrication benefits in the mixed lubrication regime. However, CS2 + κC did offer effective boundary lubrication with respect to κC, owing to the gelatinized starch + κC somehow forming a lubricating film. This highlights the crucial nature of the state of gelatinization of the starch in understanding the friction and lubrication properties of such mixtures. Ongoing studies are focusing on identifying the exact mechanisms of the tribological behavior, to allow their fine‐tuning for fat mimetic applications.In this study, we investigated the rheological and tribological properties of Kwan‐Mo You: Data curation; formal analysis; investigation; methodology; validation; visualization; writing‐original draft. Brent S. Murray: Methodology; supervision; visualization; writing‐review and editing. Anwesha Sarkar: Conceptualization; funding acquisition; methodology; project administration; supervision; visualization; writing‐review and editing.Conflict of Interest: The authors declare that they do not have any conflict of interest.Ethical Review: This study does not involve any human or animal testing.Informed Consent: Not applicable.AppendixS1: Supplementary InformationClick here for additional data file.


Triticum aestivum L.). Starch consists of amylose and amylopectin and is the major constituent of mature grain. Therefore, starch metabolism in the endosperm during grain filling can influence kernel weight. In this study, we sequenced 87 genes involved in starch metabolism from 300 wheat accessions and detected 8,141 polymorphic sites. We also characterized yield-related traits across different years in these accessions. Although the starch contents fluctuated, thousand kernel weight (TKW) showed little variation. Polymorphisms in six genes were significantly associated with TKW. These genes were located on chromosomes 2A, 2B, 4A, and 7A; none were associated with starch content or amylose content. Variations of 15 genes on chromosomes 1A and 7A formed haplotype blocks in 26 accessions. Notably, accessions with higher TKWs had more of the favorable haplotypes. We thus conclude that these haplotypes contribute additive effects to TKW.Kernel weight is a key determinant of yield in wheat ( Triticum aestivum L.) yield depends on three main components: spike number per area, grain number per spike, and thousand kernel weight (TKW). Because starch accounts for about 65–80% of the grain endosperm  and maize (Zea mays L.) (GBSSI can reduce amylose contents in rice (Oryza sativa L.), maize, barley (Hordeum vulgare L.), and wheat   . ADP-glumays L.) . Amylosend wheat . Notablyype (WT) . In addigranules . Overexpgranules . Decreasin wheat . TherefoAlthough these genes have been widely studied, systematic studies on their functional variation are lacking, especially for wheat accessions. Here, we sequenced 87 genes involved in starch metabolism, using high-throughput sequencing. We identified over 8,000 variants in 300 wheat accessions and tested them for association with thousand kernel weight (TKW), starch content (SC), and amylose content (AC), to identify useful polymorphisms among the natural populations. Notably, variants of 15 genes located on chromosomes 1A and 7A formed haplotype blocks in one subgroup. Multiple variations of genes were associated with TKW, starch weight per grain (SW), and amylose weight per grain (AW). The favorable haplotypes had positive additive effects on TKW. These results provide insights relevant to the analysis of gene function, and the variations we detected might be valuable in wheat production.Three hundred wheat accessions were used in the study: 115 Chinese landraces (CLs) and 55 modern Chinese cultivars (MCCs) from the mini-core collection , 127 forMature grains (100 g) of CL and MCC accessions were processed into flour with a Quadrumat Senior mill . The flour for each accession was then processed to quantify starch content by the AOAC Method 996.11 , using t1. Sequences were aligned with GATK2 to detect polymorphic sites relative to the reference genome of Chinese Spring (IWGSCv1.0)3.All accessions were genotyped by the MolBreeding Biotechnology Co., Ltd. . The genome-specific primers were designed by the GenoPlexs method (MolBreeding). The PCR reaction contained 50 ng DNA, 10 μL GenoPlexs 2 × G Master Mix and 8 μL primer mixture. PCR was carried out using an ABI 9700 machine with the following settings: denaturation at 95°C for 5 min, followed by 19 cycles of 95°C for 30 s, annealing at 60°C for 8 min, and extension at 72°C for 5 min. The DNA concentrations were measured by Qubit Fluorometric Quantitation , and the DNA library was sequenced by BGI . FastQC was used for quality control of the dataP of association signals was set as the 0.05/SNP marker number ; namely, -log10P > 4.343. The agronomic traits and genetic effects of favorable haplotypes in different groups were tested via ANOVA in SPSS 21.0. Pearson’s correlation coefficient was calculated between pairs of variables using the data per accession. Correlations were tested for significance using the F-test at a significance level of 1% (P < 0.01).The genetic structure of the cultivar set was evaluated by Structure 2.3.4  using 8,FST tests for the 87 genes were carried out with Arlequin 3.5.1.2  of different groups was detected for all genes in the three groups. Large differences in FST between Groups 1 and 2/3 were detected in 15 genes: 2 genes on chromosome 1A (SSIII-1A and SSIV-1A) and 13 genes on 7A , but SC did not differ significantly among the three groups. A significant difference in AC was detected only between Groups 2 and 3 in 2018 (P < 0.05). Significant differences in HD existed between Groups 1/2 and 3 (P < 0.05), but the greatest difference in HD between Groups 1 and 3 was only 4.4 days, in 2017.Thousand kernel weight (TKW), starch content (SC) and amylose content (AC), starch weight per grain (SW), amylose weight per grain (AW), and heading date (HD) of Chinese accessions from the three groups were measured in 2017 and 2018 . The traR2 = 0.0956). The SW, AW, and HD exhibited the same trends as TKW in 2017 and 2018. The TKW was correlated with SW and AW, but TKW was correlated not with SC, AC  were associated with TKW in 2017 and 2018 . The significance threshold was given as and 2018 . Associaand 2018 .FST of Group 3. Three genes associated with TKW had a different FST in Group 3 , and in the second  and eleventh  exons of SSI-7A.Because significant differences in TKW were observed between Group 1/2 and Group 3, we examined differences in 10P) of loci on two genes, SUS2-2B and SBEIIa-2A, were above 4 with AC , TaAGPSS, TaSSI, and TaSBEII (downregulated) and led to a reduction in starch and amylopectin contents  that may be associated with AC . A DH or recombinant inbred (RIL) population derived from accessions in Groups 1 and 2/3 could be used to analyze these specific variations further.Studies of SC or amylose content (AC) in cereal crops have mainly relied on functional mutational analysis and QTL mapping. In rice, an A-to-G change that results in an Asplandrace . Severalcontents . In a who 22 DAF . In the  with AC ; their f with AC . PCA sim with AC , which sSUS2-2A and SSI-7A, which form different haplotypes, could be developed into functional markers for high TKW. However, functional mutations in natural populations are limited. Several transcription factors affect starch biosynthesis in rice and maize  can be found in the article/ JH performed the experiments and data analysis and prepared the manuscript. YL contributed to collecting and measuring phenotypic data in 2017 and 2018. CH, TL, and HL provided experimental material. XZ, the corresponding author, conceived the original research and revised the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Two coronaviruses—severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV)—have caused two large-scale pandemics in the past two decades . Now, thBecause of the less cost-effective than treatment and reduce morbidity and mortality without long-lasting effects, vaccines are the most effective strategy for preventing infectious diseases . Howeverhttp://biopharm.zju.edu.cn/coviedb/).Considering the seriousness of the recent outbreaks of zoonotic coronaviruses, therapeutic agents and vaccines for pan-coronaviruses should be developed to cope with the hCoV outbreaks in the present and in the future. Here, we predict all the potential B/T cell epitopes for SARS-CoV, 2019-nCoV, and MERS-CoV to provide potential targets for pan-coronaviruses vaccine development. The prediction are based on their proteins sequences. RaTG13-CoV is included because of its high homology with 2019-nCoV (96% whole genome identity ). All thhttps://www.ncbi.nlm.nih.gov/).The protein sequences of SARS-CoV, 2019-nCoV, MERS-CoV, and RaTG13-CoV are downloaded from NCBI (The human leukocyte antigen (HLA) alleles used for T-cell epitopes prediction are derived from Zhou et al. which analyzed the HLA distribution of 20,635 individuals of Han Chinese ancestry . The topThe B-cell epitopes were predicted by the seven tools embedded in the Immune Epitope Database and Analysis Resource (IEDB) . More spAll tools give the score to define whether an amino acid is part of B-cell epitopes but not to define whether a peptide is B-cell epitopes. Here, we set B_score to quantify the possibility of a peptide to be B-cell epitopes, which is calculated as follows:L is the length of the peptide, a is the amino acid that belongs to the peptide, and an is the number of tools convinced that amino acid is part of B-cell epitopes.where The T-cell epitopes prediction were divided into two parts. One of them are presented by HLA I allele and would induce the activation of CD8+ T cells. This type of T-cell epitopes were predicted by NetMHCpan 4.0 , MHCflurIn the prediction of T-cell epitopes presented by HLA I alleles, all peptides with length range from 8 to 11 were selected and combined with previous HLA I alleles to create HLA-peptide pairs. It’s similar to predict that presented by HLA II alleles, with the difference that peptide length ranges from 15 to 28. Only HLA-peptide pairs satisfied with all thresholds of used tools would be convinced as potential T-cell epitopes in this study.orf3b is overlapped with orf3a and E gene, orf7b is overlapped with orf7a, orf8b is overlapped with orf8a, and orf9b is part of orf9a (N gene). In 2019-nCoV, only orf7b is overlapped with orf7a and other genes are separated. In MERS-CoV, the orf4b is overlapped with orf4a and orf8b is part of N gene. In RaTG13-CoV, ns7b and ns7a are overlapped.All selected coronaviruses have similar genome organization with coding genes of spike (S protein), envelope (E protein), membrane (M protein), nucleoprotein (N protein), and several open reading frames. SARS-CoV, 2019-nCoV, MERS-CoV, and RaTG13-CoV express 9, 8, 10, and 9 non-redundant protein coding genes, respectively . With four main pages “B-epitope”, “T-epitope”, “Peptide”, and “Validated”, researchers could find useful information easily and quickly. The predicted results of B-cell epitopes could be searched in “B-epitope” page. With the virus and gene selected, the corresponding predicted B-cell epitopes would appear. The predicted results of T-cell epitopes could be searched in “T-epitope” page. Similar with that in “B-epitope” page, coronavirus and protein are necessary. Besides, the type of T-cell epitopes should also be selected. Only the peptide-HLA pairs which satisfied thresholds of all tools would be showed in this page. The searchable data in the “Peptide” page is the combined result of previous predicted B-cell epitopes and T-cell epitopes. In this page, the only selectable parameter is the protein. The “Validated” page containing the predicted B/T epitopes that have been validated by recently literatures  located. So, we further investigated the shared epitopes that located in S protein. There are 265 potential epitopes in S protein shared by three coronaviruses and 35 of them with B_score larger than 5  development but also provide the targets for drug design such as neutralizing antibody on 2019-nCoV and the possible coronavirus outbreak in the future.http://biopharm.zju.edu.cn/coviedb/).The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found below: COVIEdb , the National Natural Science Foundation of China (Grant No. 31971371), the Zhejiang Provincial Natural Science Foundation of China (Grant No. LY19H300003), and the Fundamental Research Funds for the Central Universities of China.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The granule-bound starch synthase I (GBSSI) enzyme is responsible for the synthesis of amylose, and therefore, its absence results in individuals with a waxy starch phenotype in various amylaceous crops. The validation of mutation points previously associated with the waxy starch phenotype in cassava, as well as the identification of alternative mutant alleles in the GBSSI gene, can allow the development of molecular-assisted selection to introgress the waxy starch mutation into cassava breeding populations.A waxy cassava allele has been identified previously, associated with several SNPs. A particular SNP (intron 11) was used to develop SNAP markers for screening heterozygote types in cassava germplasm. Although the molecular segregation corresponds to the expected segregation at 3:1 ratio (dominant gene for the presence of amylose), the homozygotes containing the SNP associated with the waxy mutation did not show waxy phenotypes. To identify more markers, we sequenced the GBSS gene from 89 genotypes, including some that were segregated from a cross with a line carrying the known waxy allele. As a result, 17 mutations in the GBSSI gene were identified, in which only the deletion in exon 6 (MeWxEx6-del-C) was correlated with the waxy phenotype. The evaluation of mutation points by discriminant analysis of principal component analysis (DAPC) also did not completely discriminate the waxy individuals. Therefore, we developed Kompetitive Allele Specific PCR (KASP) markers that allowed discrimination between WX and wx alleles. The results demonstrated the non-existence of heterozygous individuals of the MeWxEx6-del-C deletion in the analyzed germplasm. Therefore, the deletion MeWxEx6-del-C should not be used for assisted selection in genetic backgrounds different from the original source of waxy starch. Also, the alternative SNPs identified in this study were not associated with the waxy phenotype when compared to a panel of accessions with high genetic diversity.Although the GBSSI gene can exhibit several mutations in cassava, only the deletion in exon 6 (MeWxEx6-del-C) was correlated with the waxy phenotype in the original AM206–5 source. Starch is widely used for multiple commercial purposes. Not only does it provide more than 80% of the calories in the human diet , it alsoAmylopectin, the basic unit of the starch granule, is composed of many short chains of glucose molecules. In contrast, amylose is a smaller molecule with longer chains . Starch Though it is possible to perform chemical and physical modifications of starch to change its functional properties to meet specific market demands, there are certain limitations on these modifications. In addition, regardless of the type, making such changes to starch is always associated with a higher industrial cost. Another aspect to be considered is that consumers are increasingly demanding products that are more natural, with a minimum of industrial processing. Hence, there is a growing market for naturally differentiated starches. Breeding programs can contribute to the discovery of genetic variants of interest and the further incorporation of these characteristics in commercial varieties, and waxy starch is one of these special traits.wx-D7 allele involves a 30-bp deletion at the junction of the 7th exon–intron and generates an abnormal transcript retaining the 7th intron, which introduces an immature stop codon and inactivates GBSSI. In rice, two SNPs were identified in exons 6 and 10 of the GBSSI gene, with associations to the amylose content and paste properties . Indeed, other authors have reported the great flexibility and high accuracy of the KASP technique for high-performance genotyping . The KASAnalyses in different species support the perception that the waxy phenotype is related to the absence/deficiency of the GBSSI enzyme, associated with partial and complete deletions of the gene, the presence of SNPs, and transposable elements, sometimes leading to stop codons , 32, 33.Another important aspect is the phenotyping of individuals for quantitative determination of the amylose content, which could allow the identification of alternative allelic patterns for different amylose contents. According to Dobo et al. , three pArabidopsis thaliana (L.), which is related to amylose synthesis. This protein acts on targeting and GBSSI transport  until the collection of the seeds after natural dehiscence.1 plants of each family were evaluated for the presence of waxy starch using the 2% iodine test (2 g KI and 0.2 g I2 in distilled water), applied to a cross section of the roots of all genotypes  markers.