| 摘要: |
| [目的]克隆枯草芽孢杆菌(B. subtilis)生物素基因(bioW),在bioW基因缺陷型大肠杆菌中进行表达,探讨其对bioW基因缺陷型大肠杆菌的遗传互补作用和对宿主的生长抑制作用,为生物素基因工程菌的构建奠定基础.[方法]用大肠杆菌表达载体pEXT20和枯草芽孢杆菌bioW基因构建表达载体pEXT20-biow,并将其转化到大肠杆菌DH5α和生物素基因缺陷型大肠杆菌株R876中,构建重组菌DH5α(pEXT20-bioW)和R876(pEXT20-bioW).分别将R876、DH5α(pEXT20-bioW)、R876(pEXT20-bioW)在不同培养基和不同IPTG浓度下进行培养,检测bioW基因对bioW基因缺陷型大肠杆菌株的遗传互补和生长抑制作用.[结果]在生物素限制性培养基中,只有同时加入庚二酸和IPTG时,R876(pEXT20-bioW)才能生长.在不含IPTG的LB培养基中,R876(pEXT20-bioW)生长正常;在添加不同浓度IPTG的LB培养基中,DH5α(pEXT20-bioW)和R876(pEXT20-biow)生长均受到不同程度的抑制.[结论] bioW基因对bioW基因缺陷型大肠杆菌有遗传互补作用.在大肠杆菌中,bioW基因的表达对宿主有生长抑制作用. |
| 关键词: 枯草芽孢杆菌 大肠杆菌 bloW基因 生长抑制 遗传互补 |
| DOI： |
| 分类号:Q563.7 |
| 基金项目:国家重点新产品计划? |
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| Expression of the B. subtills bioW gene in E. coli and its growth inhibition of host |
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YANG Ming-ming YANG Zhao-xia ZHANG Xi-feng WANG ling LIU Jin-ni CEN Pei-lin
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| Abstract: |
| 【Objective】 In order to construct biotin genetic engineering bacteria,the Bacillus subtilis bioW gene involved in biotin biosynthesis pathway was investigated in this study. 【Method】 The B.subtilis bioWgene was PCR amplified from B.subtilis chromosomal DNA and,the expression vector pEXT20-bioW was constructed by cloning the bioW into the E.coli expression pEXT20.Then,the pEXT20 bioW was transformed into the E.coli R876 and E.coli DH5α to examine genetic complementation respectively.【Result】 When both pimelic acid and IPTG were supplemented in the biotin-free medium,R876（pEXT20-bioW）could grow,conforming that the bioW complemented the defect of bioC.In the LB medium (without IPTG), E.coli R876（pEXT20-bioW）grew normally,however,when high concentration IPTG was added,the growth of both DH5α（pEXT20-bioW）and R876（pEXT20-bioW）was inhibited.【Conclusion】 The bioW had genetic complementation to the bacteria lacking of biotin gene and the growth of E.coli harbouring pEXT20-bioW was inhibited. |
| Key words: Bacillus subtilis Escherichia coli bioW gene growth inhibition complementation |
