Construction and identification of 2 secreted human GAD65 fragment DNA vaccines / 中南大学学报(医学版)

ABSTRACT

OBJECTIVE@#To construct and identify 2 secreted human GAD65 fragment DNA vaccines.@*METHODS@#The GAD(190-315), GAD(490-570) cDNA and hIL-2 signal peptide cDNA were linked through overlapping PCR, respectively. The fusion gene was cloned into eukaryotic expression vector pBudCE4.1. After the DNA vaccine being determined to contain the correct target nucleotide sequence, the expression of fusion proteins was detected by Western blot.@*RESULTS@#The nucleotide sequence of the cloned gene was the same as the reported sequence, and their open reading fragment was correct. The products of these DNA vaccines were expressed and secreted in eukaryotic cell using Western blot.@*CONCLUSION@#The pBudCE4.1/SGAD(190-315) and pBudCE4.1/SGAD(490-570) secreted human GAD65 fragment DNA vaccines were successfully constructed, which is a foundation for immune prevention of type 1 diabetes.