日本三角涡虫hsp70 cDNA克隆及原核表达

热休克蛋白70是热休克蛋白家族中的重要成员,它在保护生物体免受各种胁迫中发挥重要作用。由于其惊人的再生能力,淡水涡虫作为研究再生和发育的模式动物受到研究者关注。但是,有关涡虫抗逆性的分子机制却少有报道。研究采用RACE(Rapid amplification of cDNA end)技术首次从日本三角涡虫中克隆出hsp70(Djhsp70)全长cDNA序列。Djhsp70 cDNA全长2066bp,含有1947bp的开放阅读框,编码648个氨基酸,分子量71.18kD,GenBank登录号EU380241。DjHSP70的氨基酸含有真核生物HSP70家族蛋白的三个标签序列(9–16位的IDLGTTYS、199—206位的DLGGGTFD、334–339位的IVLVGG)和末端高度保守序列EEVD。经BLAST检索分析,Djhsp70的核苷酸序列和推定的氨基酸序列与目前已知HSP70家族成员高度同源。有趣的是,HSP70亲缘关系分析表明:涡虫更靠近脊椎动物,而与无脊椎动物果蝇和线虫相距较远。为了制备抗体研究DjHSP70的组织学定位,实验还成功构建了DjHSP70表达载体,在IPTG诱导下表达出约76kD的融合蛋白。Djhsp70 cDNA的克隆与表达载体的构建为下一步工作奠定了基础。

CLONING AND PROKARYOTIC EXPRESSION OF HSP70 cDNA FROM PLANARIAN DUGESIA JAPONICA

Heat shock protein 70 (HSP70) is an important member of the heat shock protein family, and it plays a key role in the process of protecting organisms from various stresses. Owing to its powerful regenerating ability, freshwater planarian has been attached high importance as model animal for the study of development and regeneration. However, little reports has addressed on stress response in planarians. In this study, the full-length hsp70 cDNA was firstly isolated and sequenced from planarian Dugesia japon...