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Construction of DNA damage response gene pprI functiondeficient and function-complementary mutants in Deinococcus radiodurans
作者姓名:GAOGuanjun  LUHuiming  HUANGLifen  HUAYuejin
作者单位:InstituteofNuclear-AgriculturalScience,ZhejiangUniversity,Hangzhou310029,China
摘    要:PprI, a DNA damage response factor from the extraordinary radioresistant bacterium Deinococcus radiodurans, plays a central regulatory role in multiple DNA damage repair. In this study, a fusion DNA fragment carrying kanamycin resistance gene with the D. radiodurans groEL promoter was cloned by PCR amplification and reversely inserted into the pprl locus in the genome of the wild-type strain R1. The resultingpprl-deficient strain, designated YR1,was very sensitive to ionizing radiation. Meanwhile, the recombinant DNA fragment was cloned into the shuttle vector pRADZ3, and resulted in plasmid pRADK with kanamycin resistance in D. radiodurans. The fragments containing complete pprl gene and 3‘-terminal deletion pprIΔ were cloned into plasmid pRADK. The resulted plasmids designated pRADKpprI and pRADKpprIΔ were then transformed to YR1. Results show that YR1 carrying pRADKpprI was able to fully restore the extreme radioresistance to the same level as the wild-type D. raiodurans R1, whereas YR1 pRADKpprIΔ failed to do so. Construction of DNA repair switch PprI function-deficient and function-complementary mutants in D. radiodurans is not only useful to elucidating the relationship between domains and functions of PprI protein, but also opens the door to the further studies of the biological functions of PprI protein in vivo.

关 键 词:PprI  基因  DNA  细菌  Deinococcus  radiodurans  突变异种  生物体  致电离辐射  生物特征
收稿时间:2 December 2004

Construction of DNA damage response genepprI function-deficient and function-complementary mutants inDeinococcus radiodurans
GAOGuanjun LUHuiming HUANGLifen HUAYuejin.Construction of DNA damage response gene pprI functiondeficient and function-complementary mutants in Deinococcus radiodurans[J].Chinese Science Bulletin,2005,50(4):311-316.
Authors:Guanjun Gao  Huiming Lu  Lifen Huang  Yuejin Hua
Institution:(1) Institute of Nuclear-Agricultural Science, Zhejiang University, 310029 Hangzhou, China
Abstract:PprI, a DNA damage response factor from the extraordinary radioresistant bacteriumDeinococcus radi-odurans, plays a central regulatory role in multiple DNA damage repair. In this study, a fusion DNA fragment carrying kanamycin resistance gene with theD. radiodurans groEL promoter was cloned by PCR amplification and reversely inserted into thepprI locus in the genome of the wild-type strain R1. The resultingpprI-deficient strain, designated YR1, was very sensitive to ionizing radiation. Meanwhile, the recombinant DNA fragment was cloned into the shuttle vector pRADZ3, and resulted in plasmid pRADK with kanamycin resistance inD. radiodurans. The fragments containing completepprI gene and 3′-terminal deletionpprI † were cloned into plasmid pRADK. The resulted plasmids designated pRADKpprI and pRADKpprI† were then transformed to YR1. Results show that YR1 carrying pRADKpprI was able to fully restore the extreme radioresistance to the same level as the wild-typeD. raiodurans Rl, whereas YR1 pRADKpprI† failed to do so. Construction of DNA repair switch PprI function-deficient and function-complementary mutants inD. radiodurans is not only useful to elucidating the relationship between domains and functions of Pprl protein, but also opens the door to the further studies of the biological functions of PprI proteinin vivo.
Keywords:Deinococcus radiodurans R1  pprI  mutants  comple- mentary    
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