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Directed Molecular Evolution of Nitrite Oxido-reductase by DNA-shuffling 被引量：1

Directed Molecular Evolution of Nitrite Oxido-reductase by DNA-shuffling
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摘要 Objective To devellop directly molecular evolution Of nitrite oxido-reductase using DNA-shuffling technique because nitrobacteria grow extremelly slow and are unable to nitrify effectively inorganic nitrogen in wastewater treatmem. Methods The norB gene coding the ntitrite oxido-reductase in nitrobacteria was cloned and sequenced. Then, directed molecular evolution of nitrite oxido-reductase was developed by DNA-shuffling of 15 norB genes from different nitrobacteria. Results After DNA-shuffling with sexual PeR and staggered extension process PCR, the sequence was differem from its parental DNA fragmems and the homology ranged from 98% to 99%. The maximum nitrification rate of the modified bacterium of X16 by DNA-shuffling was up to 42.9 mg/L.d, which was almost 10 times higher than that of its parental bacteria. Furthermore, the modified bacterium had the same characteristics of its parental bacteria of E. coli and could grow rapidly in normal cultures. Conclusion DNA-shuffling was successfully used to engineer E. coli, which had norB gene and could degrade inorganic nitrogen effectively. Objective To develop directly molecular evolution of nitrite oxido-reductase using DNA-shuffling technique because nitrobacteria grow extremely slow and are unable to nitrify effectively inorganic nitrogen in wastewater treatment. Methods The norB gene coding the nitrite oxido-reductase in nitrobacteria was cloned and sequenced. Then,directed molecular evolution of nitrite oxido-reductase was developed by DNA-shuffling of 15 norB genes from different nitrobacteria. Results After DNA-shuffling with sexual PCR and staggered extension process PCR,the sequence was different from its parental DNA fragments and the homology ranged from 98% to 99%. The maximum nitrification rate of the modified bacterium of X16 by DNA-shuffling was up to 42.9 mg/L·d,which was almost 10 times higher than that of its parental bacteria. Furthermore,the modified bacterium had the same characteristics of its parental bacteria of E. coli and could grow rapidly in normal cultures. Conclusion DNA-shuffling was successfully used to engineer E. coli,which had norB gene and could degrade inorganic nitrogen effectively.
作者 JUN-WEN LI JIN-LAI ZHENG XIN-WEI WANG MIN JIN FU-HUAN CHAO
机构地区 Institute of Environment and Health
出处 《Biomedical and Environmental Sciences》 SCIE CAS CSCD 2007年第2期113-118,共6页 生物医学与环境科学（英文版）
基金 This study was supported by the National High Technology Research and Development Program of China (863 Program) (No. 2001AA214191).
关键词 Directed evolution DNA-shuffling Nitrite oxido-reductase norB gene Nitrobacteria 亚硝酸盐氧化还原酶 DNA重排 直接分子进化 硝化细菌 废水
分类号 X172 [环境科学与工程—环境科学] Q554.4 [生物学—生物化学]
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参考文献20

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同被引文献13

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Biomedical and Environmental Sciences
2007年 第2期

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