斯氏艾美耳球虫热休克蛋白70基因的克隆及表达分析

发布时间：2018-06-06 11:24

  本文选题：斯氏艾美耳球虫 + 热休克蛋白(HSP)　； 参考：《黑龙江畜牧兽医》2017年07期

【摘要】：为了获得斯氏艾美耳球虫(E.stiedai)的热休克蛋白70(HSP70)的全基因序列,并诱导表达重组HSP70蛋白,试验根据免疫蛋白组学研究的质谱结果获得的部分氨基酸序列和已发布近源物种HSP70蛋白序列设计引物,通过c DNA末端快速扩增法(RACE)获得E.stiedai HSP70的全基因序列,将其连入原核表达载体p ET-28a(+),转化Competent Rosetta2(DE3)p Lys S,通过IPTG诱导表达重组HSP70蛋白,并进行Western-blot分析。结果表明:HSP70序列全长为2 727 bp,最大ORF为2 010 bp。HSP70在诱导后37℃培养4 h条件下有表达,但全部为不溶性表达;在诱导后20℃过夜培养条件下无表达。说明试验成功获得E.stiedai HSP70的全基因序列以及重组HSP70蛋白。
[Abstract]:In order to obtain the whole gene sequence of heat shock protein 70 (HSP70) of E. stiedai, and to induce the expression of recombinant HSP70 protein, A primer was designed based on the results of mass spectrometry and HSP70 protein sequence of the published near-source species. The whole gene sequence of E.stiedai HSP70 was obtained by rapid amplification of c DNA terminal. The recombinant HSP70 protein was transfected into prokaryotic expression vector pET-28a (pET-28a) and transformed into Competent Rosetta2(DE3)p Lys S.The recombinant HSP70 protein was induced by IPTG and analyzed by Western-blot. The results showed that the full length of the 1: HSP70 sequence was 2 727 BP, and the maximum ORF was 2 010 bp.HSP70, which was expressed at 37 鈩，

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