本文選題：牛支原體　切入點(diǎn)：P蛋白　出處：《中國病原生物學(xué)雜志》2017年01期

【摘要】：目的構(gòu)建3種牛支原體P81蛋白原核表達(dá)質(zhì)粒,分析其在大腸埃希菌中的表達(dá)情況,并純化P81蛋白。方法根據(jù)GenBank發(fā)表的牛支原體P81基因序列(GenBank登錄號(hào):AY627040.1),密碼子優(yōu)化后合成P81全基因序列。構(gòu)建原核表達(dá)質(zhì)粒pET28a-P81、pET32a-P81和pGEX-6P-1-P81,經(jīng)IPTG誘導(dǎo)后采用SDS-PAGE電泳分析P81蛋白在原核表達(dá)系統(tǒng)中的表達(dá)情況;應(yīng)用Ni-NTA對(duì)目的蛋白進(jìn)行純化。結(jié)果pET28a-P81、pET32a-P81和pGEX-6P-1-P81均高效表達(dá)分子質(zhì)量單位為78ku左右的P81蛋白。表達(dá)產(chǎn)物進(jìn)行Ni-NTA親和層析,得到純化的P81蛋白。結(jié)論構(gòu)建的重組原核表達(dá)質(zhì)粒pET28a-P81、pET32a-P81、pGEX-6P-1-P81能在大腸埃希菌中高效表達(dá)牛支原體P81蛋白,為進(jìn)一步分析該蛋白的抗原性奠定了基礎(chǔ)。
[Abstract]:Objective to construct three prokaryotic expression plasmids of Mycoplasma bovis P81 protein, analyze its expression in Escherichia coli, and purify P81 protein.Methods according to the P81 gene sequence of Mycoplasma bovis published by GenBank and GenBank accession number: AY627040.1, the whole P81 gene sequence was synthesized after the codon was optimized.The prokaryotic expression plasmids pET28a-P81, pGEX-6P-1-P81 and pGEX-6P-1-P81 were constructed. The expression of P81 protein in prokaryotic expression system was analyzed by SDS-PAGE electrophoresis after induction by IPTG, and the target protein was purified by Ni-NTA.Results both pET28a-P81 pET32a-P81 and pGEX-6P-1-P81 expressed P81 protein with 78ku or so.The purified P81 protein was obtained by Ni-NTA affinity chromatography.Conclusion the recombinant prokaryotic expression plasmid pET28a-P81 pET32a-P81 pGEX-6P-1-P81 can efficiently express Mycoplasma Bovis P81 protein in Escherichia coli, which lays a foundation for further analysis of the antigenicity of the protein.
【作者單位】： 軍事醫(yī)學(xué)科學(xué)院軍事獸醫(yī)研究所;廣西大學(xué)動(dòng)物科學(xué)技術(shù)學(xué)院;吉林農(nóng)業(yè)大學(xué)動(dòng)物科技學(xué)院;吉林大學(xué)動(dòng)物醫(yī)學(xué)學(xué)院;延邊大學(xué);
【基金】：吉林省科技發(fā)展計(jì)劃項(xiàng)目(No.20140309024NY,20140623019TC,20160623024TC)
【分類號(hào)】：S852.62
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本文編號(hào)：1720698