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缩短5′-UTR序列可以提高hGH基因在昆虫细胞中的表达
Enhancing hGH Expression Level in Insect Cells by Shortening the 5′-UTR of hGH cDNA
【Abstract】 The regulation of foreign gene expression in Insect-Baculovirus Expression System is very complex. In this report, the effect of 5′-UTR in the expression of hGH gene in cultured Sf9 cells was examined. A 18 bp length in the end of 5′-UTR of hGH (human Growth Hormone, hGH) cDNA including a stem-loop structure was deleted by PCR. The truncated hGH cDNA, Δ1hGH was cloned in pFastBac1, named pFast-Bac-Δ1hGH. After transforming into E.coli. DH10Bac, which have a shuttle vetor-Bacmid, the Δ1hGH was integrated into Bacmid by site-specific transposition, and an expression vector, rBacmid-Δ1hGH DNA was acquired. By transfecting the cultured Sf9 cells with the recombinant expression vector DNA, pure recombinant virus, rAcV-Bac-Δ1hGH was obtained, and hGH gene was expressed. Immuno-blot and Chemiluminescent assay revealed that the expressed hGH had normal immunological activity, the ammount of hGH expression level in Sf9 cell supernatant infected with rAcV-Bac-Δ1hGH containing the truncated 5′UTR was four to five times higher than that infected with rAcV-Bac-hGH.
【Key words】 hGH gene; PCR; 5′-UTR; Bac-to-Bac; expression;
- 【文献出处】 生物工程学报 ,Chinese Journal of Biotechnology , 编辑部邮箱 ,2002年04期
- 【分类号】Q786
- 【被引频次】2
- 【下载频次】74