Cloning Expression in E.coli and Biological Activity of Human Thymosin β₄

CHE Yan-Ke¹, YANG Hui, LU Fan, PU Qin, LI Ren-De¹, ZHAO Zhong-Liang^*
( Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi'an 710032, China; ¹College of Life Sciences, Lanzhou University, Lanzhou 730000, China )

Abstract    The cDNA thymosin β₄ was synthesized by combining of chemical and enzymatic methods. First, two complement fragments of thymosin β₄ cDNA were synthesized by DNA synthesizer, and then denatured, annealed and extended by DNA polymerase. This fragment of thymosin β₄ was then inserted into the EcoRV and HindIII restriction endonuclease site of an expression plasmid pLDH4 (a kind of E.coli plasmid) by blunt and cohesive ligations. Finally, the recombinant plasmid which expressed thymosin β₄ was screened by digestion and DNA sequencing. This recombinant plasmid highly expressed the thymosin β₄, which accounted for 30% of total bacteria proteins. By salting out and chromatography, a 95% purity of recombinant thymosin β₄ was obtained. Biological assay indicated that the recombinant thymosin β₄ could induce lymphocyte proliferation and differentiation.
Key words    thymosin β₄; gene expression; E-rose; MTT

^*Corresponding author： Tel, 86-29-3374537; e-mail, [email protected]