Full-Lengh Cloning Of HCMV UL49 Gene And Bioinformatical Analysis

Objective: To obtain the full lengh cDNA of HCMV UL49 gene by SMART Rapid Amplification of cDNA Ends technology in HCMV AD169. Using bioinformatics methods to analyse UL49 gene's function. Setting up foundation for further research of UL49 gene.Methods: Inoculate HCMV AD169 to well growth MRC-5 cell; collect the cells while the MRC-5 cell appear cytopathic effect; Extract HCMV RNA from cells, After obtained 5'RACE and 3'RACE of cDNA of HCMV UL49 by SMART Rapid Amplification of cDNA Ends technology. The UL49 genes primers were designed according to the result of RACE. Amplified full lengh cDNA of HCMV UL49 gene by RT-PCR. Then cloned into pMD18-T simple vector and sequencd subsequently; Use bioinformatics methods to analyse UL49 gene's biological function.Results: We had effected AD169 to well growth MRC-5 cell successfully. Using PCR, we detected the special gene for AD169: IEA and LA gene. Using Smart assay, we abtained and cloned the 3' and 5'termination of UL49 gene. Sequencing results suggested its 3'-untranslated region and 5'-untranslated region were 315bp and 90bp seperatly. The full length cDNA of UL49 gene, 2118bp, was amplified using RT-PCR. Bioinformatical analysis suggested UL49 protein was a transmembrane protein.Conclusion: We had obtained the full lengh cDNA of HCMV UL49 gene by SMART Rapid Amplification of cDNA Ends technology. The cDNA sequence is 2118bp in size, The 5'-untranslated region is 90bp, while the 3'-untranslated region is 315bp.the open reading frame is 1710bp and capable of encoding 570 amino acids. It's an unknow protein. The result of the bioinformatics analysed indicated UL49 is a transmembrane protein.