NT4-Al融合基因原核表达载体的构建

目的构建神经营养素（NT4）与肿瘤血管抑制肽Alphastatin融合基因的原核表达载体pBV220-NT4-Al。方法采用非对称互补引物（模板）法，依据GenBank提供的Alphastatin基因序列，设计合成并扩增出肿瘤血管抑制肽Alphastatin的cDNA；将其连接到NT4的信号肽和前导序列3’端，组成融合基因NT4-Al，再将该融合基因亚克隆于表达载体pBV220，构建pBV220-NT4-Al。结果经DNA测序、限制性内切酶酶切等方法证实Alphastatin成功重组到NT4信号肽和前导序列3’端，并将融合基因亚克隆于pBV220内。结论成功构建原核表达载体pBV220-NT4-Al，为进一步研究肿瘤基因治疗奠定了基础。

Construction of prokaryotic expression vector for NT4-Al fusion gene

Objective To construct the prokaryotic expression vector, pBV220-NT4-Al, by fusing genes neurotrophin 4 （NT4） and alphastatin （A1）. Methods According to Alphastatin gene sequence in Gene Bank, double stranded cDNA of Alphastatin was gained by asymmetrical primer/template. Alphastatin cDNA was ligated to the signal peptide and 3＇ leader sequence of neurotrophin 4 （NT4）. Then it was subcloned into prokaryotic expression vector pBV220, and forming pBV220-NT4-Al. Results Evidences of DNA sequence analysis and restriction enzymes digestion showed that the Alphastatin cDNA had already combined to the 3＇ terminal of the signal and leader peptides of NT4, and the fusion gene was subcloned into pBV220 successfully. Conclusion Prokaryotic expression vector pBV220-NT4-Al is constructed successfully, which lays a foundation for further study of tumor genetic therapy.