目的 构建表达重组胸腺素α1(Tα1)的pMAL- C2x - Tα1/TBI工程菌.方法 将人工合成的Tα1序列进行PCR扩增,将扩增的片段和pMAL- C2x质粒载体分别经BamHI和EcoR I双酶切后,用T4 DNA快速连接酶连接构建pMAL- C2x - Tα1融合表达质粒,再经测序正确后,将重组体转化至大肠埃希菌TB1菌中,pMAL- C2x -Tα1/TB1菌在LB液体培养基中培养,经IPTG诱导表达麦芽糖结合蛋白与Tα1的融合蛋白(MBP- Tα1),采用Westernblot对MBP- Tα1进行鉴定.结果 pMAL-C2x-Tα1/TB1工程菌能有效表达MBP- Tα1,融合蛋白占菌体蛋白的33.6％,分子量约为45×103.结论 工程菌的成功构建和表达为重组Tα1的纯化、生物学活性等研究奠定了基础.%Objective To construct an engineering strain of pMAL- C2x - Tal/TBl which can express Thymosin al (Tα1). Methods Tα1 gene was synthesized and amplified by PCR, PCR product and pMAL- C2x vector were digested with restriction endonuclease BamHI and EcoR I respectively, and were linked with T4 DNA ligase to construct pMAL- C2x -Tal. After being sequenced, pMAL- C2x - Tα1 vector was transformed into E. coli TB1 for fusion expression under induction of IPTG. . The expressed product was identified by Western blot. Results The DNA sequence of the synthesized Tal gene was identical to the original design. The constructed recombinant plasmid pMAL- C2x - Tα1 was highly expressed in E. coli TB1. The BMP - Tal fusion protein was about 33. 6% of the total bacteria protein and the relative molecular weight of BMP - Tal fusion protein was about 45 x 103. Conclusion The successful construction and expression of pMAL-C2x - Tal/TBl laid a foundation for the research of purification and biological acitivity of recombinant Tα1.
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