本文選題：腦心肌炎病毒 + VP基因�。� 參考：《中國獸醫(yī)學(xué)報》2017年04期

【摘要】：應(yīng)用RT-PCR技術(shù)擴增腦心肌炎病毒(EMCV)BD2株VP1基因,將其克隆至原核表達(dá)載體pET-32a(+),利用在線分析軟件對該基因所編碼的蛋白序列進(jìn)行生物信息學(xué)分析。將重組質(zhì)粒pET-32a-VP1轉(zhuǎn)化到E.coli BL21(DE3)感受態(tài)細(xì)胞中,在不同濃度的IPTG和不同溫度條件下對目的蛋白進(jìn)行誘導(dǎo)表達(dá)。結(jié)果顯示,EMCV VP1基因全長831bp,編碼277個氨基酸,VP1蛋白為酸性、親水性蛋白質(zhì),蛋白空間結(jié)構(gòu)以α-螺旋、β-折疊和無規(guī)則卷曲為主。經(jīng)SDS-PAGE分析可知,VP1蛋白在37℃,1.0mmol/L IPTG的誘導(dǎo)條件下以包涵體形式獲得了高效表達(dá),重組蛋白的相對分子質(zhì)量約為49 300。Western blot結(jié)果表明其具有良好的反應(yīng)原性。綜上所述,EMCV VP1蛋白是一種抗原性較高的親水性蛋白,在原核系統(tǒng)中能高效表達(dá),該研究為進(jìn)一步建立相應(yīng)的抗體檢測方法和DNA疫苗的研制提供理論基礎(chǔ)。
[Abstract]:The RT-PCR technique was used to amplify the VP1 gene of encephalitis virus strain EMCVV BD2, and cloned it into the prokaryotic expression vector pET-32a (pET-32a). The protein sequence encoded by the gene was analyzed by bioinformatics using on-line analysis software. The recombinant plasmid pET-32a-VP1 was transformed into E.coli BL21 (DE3) competent cells, and the target protein was induced to express under different concentrations of IPTG and different temperature. The results showed that the total length of EMCV VP1 gene was 831 BP, encoding 277 amino acid protein VP1 as acidic and hydrophilic protein. The spatial structure of the protein was 偽 -helix, 尾 -fold and irregular curl. SDS-PAGE analysis showed that the VP1 protein was highly expressed in the form of inclusion body under the induction of 1.0 mmol / L IPTG at 37 鈩，

本文編號：1915583