Swine major histocompatibility complex (SLA) class II DMB and LMP2 are two vital antigen presenting related functional genes. Through the cDNA clones of the genes and the screening of their variance, some important information and gene data could help to research the mechanism of antigen presenting in immune response and develop the swine immunogenetics and breeding for disease resistance. In this research, we used the EST Blast to in silico clone novel pig genes. Moreover, we predicted the function of the putative peptides encoded by the genes. The aim of this study is to establish available cloning gene approach based the high similarity of homologous gene sequences in different species.1. Cloning and functional analysis of SLA-DMB geneA novel pig gene (SLA-DMB) full-length cDNA of 1456bp was cloned by Blasting the pig EST database with the human homologous DMB gene cDNA sequence information as a querying probe. Searching the cDNA sequence for potential coding regions by ORF finder (NCBI), an entire open reading frame (ORF) of 263 amino acids was detected with a potential start-codon at the 277th site and a stop codon at the 1068th site. To examine the accuracy of in silico cloning cDNA sequence, we cloned SLA-DMB gene by RT-PCR and sequenced a cDNA fragment covering the coding region of it (GenBank Accession, DQ431246). The result confirmed that the in silico cloning cDNA sequence was very accurate. The results of the multiple sequences alignment and phylogenetic analyses revealed the putative peptide encoded by pig SLA-DMB gene is complete, and SLA-DMB is actual functional gene in pig. The gene was analyzed for PI, Mw, Amino acid content, hydrophobicicy, protein secondary structure, protein tertiary structure and superfamily by bioinformatics methods. According to the above analysis results, we can predict the deduced encoding protein by SLA-DMB is similar function to DMBs of other species, and SLA-DMB gene plays a vital role of SLA class-â…¡ antigen presenting in pig immune system.2. The study of genotyping methodology on SSCP and variance inSLA-DM genesThe polymorphism of the third exon about SLA-DMA was studied in tibetan pigs from Ganzi municipality. The complicated electrophoresis patterns based on single strand conformation polymorphism (SSCP) was discussed by combining LIS-SSCP, molecular clone sequencing, bacterical colony PCR, asymmetric PCR and structure prediction of DNA single-strand. There was great identity between the genotyping of SSCP and the structure predictions of DNA single-strand. Hence, a rapid and exact method was established and developed its application based on genotyping of PCR-SSCP. We scanned... |