摘要
目的表达βＢ２－晶体蛋白，以研究其结构与功能的关系。方法用ＲＴ－ＰＣＲ的方法从大鼠晶状体中获得βＢ２－晶体蛋白的ｃＤＮＡ，将ｃＤＮＡ克隆到表达载体ｐＭＯＮ５７４３，转染Ｅ．ｃｏｌｉ菌株ＪＭ１０１，以萘啶酮酸诱导表达βＢ２－晶体蛋白，用ＤＥＡＥ纤维素层析纯化。结果ＲＴ－ＰＣＲ扩增获得约７００ｂｐ的ｃＤＮＡ，在Ｅ．ｃｏｌｉ中获得高效表达，βＢ２－晶体蛋白占细菌总蛋白的３０％。ＤＥＡＥ纯化后获一２４ｋＤ蛋白，分子量与βＢ２－晶体蛋白一致。结论βＢ２－晶体蛋白ｃＤＮＡ可在大肠杆菌中获高效表达，表达产物的分子量约２４ｋＤ。
Objective To obtain recombinant βB2-crystallin for the study of its structure-function relationship.Methods The cDNA encoding βB2-crystallin was obtained from rat lenses by RT-PCR, cloned into expression vector pMON5743 and introduced into the competent cells of E.coli strain JM101. βB2-crystallin expression was induced by the addition of nalidixic acid and the expressed protein was purified from E.coli cells using ion-exchange chromatography with DEAE DE52 cellulose. Results A cDNA of βB2-crystallin,of about 700 bp, was obtained by RT-PCR. βB2-crystallin was expressed at high level and the yield of βB2-crystallin in E.coli was about 30% of the total cellular protein. After purification, a 24kD protein was obtained, which is consistent with the molecular weight of βB2-crystallin.Conclusion High level expression of βB2-crystallin can be obtained in E.coli. The molecular mass of recombinant βB2-crystallin is about 24kD
出处
《徐州医学院学报》
CAS
1998年第6期444-446,共3页
Acta Academiae Medicinae Xuzhou
