果蝇nulp1特异性多克隆抗体制备及检测

Polyclonal Antibody Preparation of Nulp1 Gene in Drosophila

为了利用果蝇模型研究nulp1蛋白在早期心脏发育中的功能,采用PCR技术扩增出果蝇nulp1基因的部分编码区并插入到pET28a表达载体中.之后将重组质粒转入大肠杆菌(E.coli)Rosetta(DE3);通过IPTG诱导表达His-nulp1融合蛋白,该融合蛋白采用Ni2+金属螯合层析纯化后,免疫新西兰兔制备了多克隆抗体,利用west-ern blot对抗体进行分析.结果表明获得了高效价的特异性兔抗f-nulp1多克隆抗体.这为进一步研究果蝇心脏发育的分子机制创造了条件.

To understand the function of the nulpl protein in early heart development by using Drosophila as a model, here, a fraction of the encoded region in the Drosophila nulpl gene was obtained by PCR amplification, and then was inserted into pET28a vector to establish the expressing system. Then, the recombinant plasmid was transformed into Rosetta （DE3）, and the fusion protein His-nulpl was induced by IPTG. After purification by Ni2＋ metal chelate chromatography, His-nulpl fusion protein was used to immune the New Zealand white rabbits to prepare polyclonal antibody. The antibody was assayed by western blotting and immunohistochemistry. The result show that the polyclonal antibody is of high sensitivity and specificity. The antibody will facilitate further studies on the molecular mechanism of heart development in Drosophila.