果蝇Mef2基因多克隆抗体的制备及检测

Preparation and Detection of Polyclonal Antibody of Mef2 Gene in Drosophila

为了在果蝇模型中进一步研究Mef2基因的功能,制备了Mef2多克隆抗体.以果蝇cDNA文库为模板,PCR扩增出亲水性和特异性均好的果蝇Mef2基因片段,将其克隆入pET-28a原核表达载体,然后将重组质粒转化入大肠杆菌菌株Rossetea,用IPTG诱导表达融合蛋白.融合蛋白先经Ni-IDA凝胶柱纯化,再免疫新西兰大白兔制备多克隆抗体,然后用不同浓度的抗体分别进行Western-blot实验检测抗体效价.所得结果显示获得了较高效价的果蝇Mef2多克隆抗体.

In order to study the further function of Mef2 gene in Drosophila model, it is necessary to prepare its polyclonal antibody. The cDNA fragment of part coding sequence of Mef2 was obtained by PCR amplification, then it was cloned int6 the expression vector pET-28a. The recombinant expression plasmid was transformed into Rosseta and induced by IPTG to acquire fusion protein. The purity protein obtained by treating the lysates with Ni- IDA gel column purification, then it was immuned into the New Zealand white rabbit. The antibody＇s fiter and specification was identified by Western blot separately, and all the results show that high sensitivity and specificity anti- Mef2 polyclonaI antibody was generated.