Sequence analysis of the full-length genome of Japanese encephalitis virus vaccine SA_(14)-14-2 strain
Objective To sequence the full length genome of Japanese encephalitis virus attenuated live vaccine SA 14 14 2 strain, provide direct information about the genomic structure and the possible relationship to the attenuated mechanism, and set up the basis for investigating the genetic quality control markers of the vaccine. Methods Six pairs of overlaping primers were designed according to the published sequences of SA 14 14 2 and SA 14 strains. Using RT PCR, cDNA fragments of SA 14 14 2 strain were got and cloned into pGEM T vector, then transformed into competent TG1 hosts. Positive clones were screened, identified and sequenced. Results Sequence analysis showed that genome of JEV vaccine SA 14 14 2 strain consisted of 10976 nucleotides and contained a single open reading frame of 10299 which encoded a polyprotein of 3432 amino acids. Compared with the published sequence of SA 14 14 2 and SA 14 strain, the homology of the nucleotide sequence and deduced amino acid sequence were all more than 99%, the mutation sites were located in different regions. Seven mutation sites responsible for the attenuation deduced were all consisted in. Among them, there were 4 mutated sites between 1296 and 1506 in the E region, this region may be used as genetic markers for the quality control of the vaccine. Conclusion Sequence of SA 14 14 2 vaccine genome is similar to the published data. Some mutations are among them probably due to passage of virus in difference cultures. The sequence analysis would be helpful to understanding attenuated mechanism of the vaccine and developing genetic quality control standard for vaccine production.