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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

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【作者】 耿朝晖刘莹郜鹏赵东明李澍俞新大张宝珠

【Author】 GENG Zhao-Hui LIU Ying GAO Peng ZHAO Dong-Ming LI Shu YU Xin-Da\+* ZHANG Bao-Zhu (Department of Biochemistry & Molecular Biology, Nankai University, Tianjin 300071, China)

【机构】 南开大学生物化学与分子生物学系南开大学生物化学与分子生物学系 天津300071天津300071天津300071

【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.

【关键词】 hGH基因PCR5′-UTRBac-to-Bac表达效率
【Key words】 hGH genePCR5′-UTRBac-to-Bacexpression
【基金】 国家“九五”重点科技攻关项目 (No .96 12 0 46 0 1)基金资助~~
  • 【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
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