编码日本血吸虫Akt蛋白的cDNA克隆及原核表达

CLONING AND PROKARYOTIC EXPRESSION OF A CDNA ENCODING AKT PROTEIN OF SCHISTOSOMA JAPONICUM

本研究利用生物信息学分析日本血吸虫编码Akt蛋白的cDNA并对编码该蛋白的cDNA进行了克隆和原核表达。生物信息学分析表明日本血吸虫存在两个Akt蛋白。利用PCR将其中一个Akt蛋白催化功能域的编码cDNA进行了克隆,并利用原核表达系统对此蛋白片段进行诱导表达并进行了重组蛋白的纯化,将重组蛋白免疫新西兰大白兔制备了多克隆抗体。Western blot分析表明,该抗体能被日本血吸虫Akt重组蛋白特异性识别。本研究为进一步研究该蛋白的功能奠定了基础。

In the study, we employed bioinformatics and molecular techniques to preliminary study the cDNAs encoding for Akt proteins in Schistosoma japonicum. Bioinformatical analysis indicated that Schistosoma japonicum has two Akt proteins. Subsequently, the cDNA fragment encoding the catalytic domain of a Akt protein was amplified by PCR technique and then cloned into a expression vector pET28a（＋） . The expressed recombinant protein was purified by using a Nickel charged affinity column. The polyclonal antibodies against the Akt protein were raised by immunizing the purified protein in Newland rabbit. Western blot analysis indicated that the polyclonal antibodies were able to specifically recognize the recombinant SjAkt protein. Overall, the present study provides fundamental information for further investigating the functions of Akt proteins in Schistosomajaponicum.