柔嫩艾美耳球虫杨凌株TA4基因的克隆和原核表达

根据国外报道的序列设计1对引物,以纯化的柔嫩艾美耳球虫(E.tenella)杨凌(YL)株孢子化卵囊的总RNA为模板,用RT-PCR方法扩增出E.tenella YL株的TA4基因.序列分析表明,该序列全长741 bp,开放阅读框(ORF)为651 bp,共编码217个氨基酸,与E.tenella HT株、E.tenella BJ株和E.tenella ZJ株的TA4基因ORF序列同源性分别为99.8%、99.8%和99.27%.由此确定所获得的目的基因为E.tenella YL株TA4基因.利用生物信息学和分子生物学软件对TA4基因编码的蛋白进行结构预测,结果表明,该蛋白为一结构松散的球状蛋白.将TA4基因克隆到表达载体pET-32a中,构建重组质粒pET-TA4并在大肠杆菌BL21中进行表达,表达产物的SDS-PAGE分析结果表明,成功地表达了分子质量为41 ku的融合蛋白.

Cloning and prokaryotic expression of TA4 gene from E.tenella YL strain

A pair of primers was designed according to TA4 gene sequence of Eimeria tenella HT strain available in GenBank.Then a TA4 gene was amplified using total RNA from the purified E.tenella YL strain sporulated oocysts as template by RT-PCR.Sequence analysis showed that ORF of TA4 gene was consisted of 651 nucleotides and encoded 217 amino acids.Compared with E.tenella HT strain,E.tenella BJ and E.tenella ZJ strain,TA4 gene from E.tenella YL strain shared 99.8%,99.8% and 99.27% homology,respectively.The protein...