来航鸡FOXL2基因的真核表达及纯化

Eukaryotic expression and purification of FOXL2 gene in leghorn

【目的】对来航鸡FOXL2基因进行真核表达及纯化,为其生物学功能研究奠定基础。【方法】根据GenBank已发表的来航鸡FOXL2基因序列(GenBank登录号:JF_708868.1)设计引物,以来航鸡全血DNA为模板,PCR扩增FOXL2基因,经EcoRⅠ和XbaⅠ双酶切后定向克隆于pPICZαA中,获得pPICZαA-FOXL2真核表达载体。将pPICZαA-FOXL2转化毕赤酵母菌GS115,用甲醇进行诱导表达,对表达产物进行SDS-PAGE和Western blotting检测。【结果】克隆了918bp的FOXL2基因;成功构建了pPICZαA-FOXL2真核表达载体;实现了FOXL2基因在毕赤酵母菌GS115中的融合表达,获得了分子质量约为37ku的重组蛋白FOXL2,经Western blotting检测其具有较好的生物学活性。【结论】成功地在毕赤酵母中表达了来航鸡FOXL2基因。

[Objective] Eukaryotic expression and purification of leghorn FOXL2 gene were conducted to understand its biological function. [Method] A pair of primers were designed based on the published nudeotide sequence of leghorn FOXL2 gene （GenBank accession number： JF_708868. 1）. The full-length coding sequence of the gene was amplified by PCR using the whole blood DNA as template. The product was double-digested by EcoR I and Xba I before being directionally cloned into the pPICZαA vector to ob- tain the eukaryotic expression vector pPICZαA-FOXL2. The recombinant expression vector was induced with methanol in GS115 and SDS-PAGE and Western blotting were used to detect the expressed protein. [Result] The FOXL2 gene of 918 bp was cloned and its eukaryotic expression vector pPICZαA-FOXL2 was successfully obtained. The FOXL2 gene fusion expression in yeast GS115 was conducted and the 37 ku FOXL2 protein was obtained,which had good biological activities as shown by Western blotting detection. [Conclusion] The leghorn FOXL2 gene was successfully expressed.