Procaryotic Expression Of Inserted Alternative Splicing Variants In Equine Lentivirus Receptor-1

Equine lentivirus receptor-1 (ELR1) is supposed to be the only recepetor of EIAV. In recent studies, it was found that the ELR1 species (termed as INR) contains an insertion of 153bp between the 786-787nt of the published ELR1 cDNA sequence. After inserted into an eukaryotic expression vector, the inserted receptor was expressed as a soluble protein of 38 ku(sELR1), and this protein could inhibit EIAV infection at the entry stage.The purpose of this experiment was to express high level of soluble receptor (sELR1) by using different prokaryotic expression systems. At First, it was used for the same enzyme sites BamH I and Not I of pGEX-6p-1and pET-30a to establish the recombinant plasmids pGEX-INR and pET-30a-INR successly. Then the induced time of transformed bacteria were Changed from 2h to 12h, and a series of the revulsant concentration of IPTG were set up, including 0.2, 0.4, 0.6 ,0.8 , 1.0mmol/L. In addition, the expression of sELR1 was induced in the different temperatures of 28℃and 37℃, or with 1%,3% sorbitol and glucose. The results showed that the vector of pET-30a-INR could express part of soluble protein in 0.2 mmol/L IPTG, 28℃, 1% sorbitol, and cultured for 10h. Through western Blot testing, the sELR1 expressing of pGEX-INR and pET-30a-INR were correct, whose molecular weight were 47Ku and 36Ku. The results was consistent with the expected. This research lays foundation for the development of the polyclonal antibody.