人aFGF cDNA的次级克隆及其重组产物的纯化

Secondary cloning of human aFGF cDNA and purificationof recombinant aFGF

目的：为使ａＦＧＦ表达产量提高，以便于纯化，将载有人酸性成纤维细胞生长因子（ａＦＧＦ）全编码区ｃＤＮＡ的ＰＫＫ２２３－３质粒自ＤＨ５α细菌转入ＪＭ１０５细菌。方法：采用超声法和溶菌酶破碎法裂解扩增的ＪＭ１０５细菌，通过ＨｅｐａｒｉｎＳｅｐｈａｒｏｓｅＣＬ－６Ｂ亲和层析法提纯表达的重组产物。结果：ＳＤＳ－ＰＡＧＥ的结果表明，纯化重组产物的分子量约为１８０００。

Objective:To increase the output of expressed recombinane aFGF so as to purify it.Methods:PKK223-3 plasmid carrying the whole encoding region cDNAof human acid fibroblast growth factor (aFGF) was subcloned from DH5α host cells to JM105 host cells.Amplified JM105 host cells were lysed by either aupersonic or lysozyme breaking methods.Expressed recombinant aFGFwas purified by Heparin Sepharose CL-6B affinity chromatography.Results:Purified protein showed a single band with a molecular weight of 18 000 in SD-PAGE analysis.Conclusion: