Isolation, analysis and prokaryotic expression of nitric oxide associated factor gene-NOA1 from potato (Solanum tuberosum L.)

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	Isolation, analysis and prokaryotic expression of nitric oxide associated factor gene-NOA1 from potato (Solanum tuberosum L.)
	Zhang, B. O.1,2; Wang, Haiqing 1; Shen, Yuhu 1; Zhang, Huaigang 1
	2010-05-03
发表期刊	AFRICAN JOURNAL OF BIOTECHNOLOGY
ISSN	1684-5315
卷号	9 期号:18 页码:2606-2612
文章类型	Article
摘要	In this study, the cDNA of a NOA (nitric oxide associated factor) was isolated with the strategy of EST in silico cloning in combination with RT-PCR and 3'RACE from potato (Solanum tuberosum L.) for the first time (named StNOA1). The StNOA1 bears a centrally positioned GTPase-binding domain. Sequence alignment and phylogenetic analysis of the deduced StNOA1 protein with other known NOA family protein indicates that StNOA1 is highly homologenous with NbNOA1 (NOA from Nicotiana benthamiana). The cDNA was cloned into prokaryotic expression vector, pET-30a (+) and expressed in Escherichia coli BL21 (DE3) after induction with IPTG. The recombinant protein was dissolved by 8 M urea and recovered by dialysis due to most of them were in inclusion bodies. Then the recovered recombinant protein was purified by Ni-NTA and analyzed by SDS-PAGE. The results of SDS-PAGE showed that the StNOA1 was successfully expressed with the pET prokaryotic expression system and purified. The present study is the basis for further elucidating the biochemical characteristics of StNOA1 and is very significant for elucidating the nature of plant NOA and its action mechanisms in endogenous NO synthesis in plant species.; In this study, the cDNA of a NOA (nitric oxide associated factor) was isolated with the strategy of EST in silico cloning in combination with RT-PCR and 3'RACE from potato (Solanum tuberosum L.) for the first time (named StNOA1). The StNOA1 bears a centrally positioned GTPase-binding domain. Sequence alignment and phylogenetic analysis of the deduced StNOA1 protein with other known NOA family protein indicates that StNOA1 is highly homologenous with NbNOA1 (NOA from Nicotiana benthamiana). The cDNA was cloned into prokaryotic expression vector, pET-30a (+) and expressed in Escherichia coli BL21 (DE3) after induction with IPTG. The recombinant protein was dissolved by 8 M urea and recovered by dialysis due to most of them were in inclusion bodies. Then the recovered recombinant protein was purified by Ni-NTA and analyzed by SDS-PAGE. The results of SDS-PAGE showed that the StNOA1 was successfully expressed with the pET prokaryotic expression system and purified. The present study is the basis for further elucidating the biochemical characteristics of StNOA1 and is very significant for elucidating the nature of plant NOA and its action mechanisms in endogenous NO synthesis in plant species.
关键词	Nos SoL.num Tuberosum L. Atnoa1
WOS标题词	Science & Technology ; Life Sciences & Biomedicine
关键词[WOS]	STOMATAL CLOSURE ; 2 ECOTYPES ; SYNTHASE ; ARABIDOPSIS ; PLANTS ; RESPONSES ; SIGNAL ; INVOLVEMENT ; RESISTANCE ; PROTECTS
收录类别	SCI
语种	英语
WOS研究方向	Biotechnology & Applied Microbiology
WOS类目	Biotechnology & Applied Microbiology
WOS记录号	WOS:000277847400003
引用统计
文献类型	期刊论文
条目标识符	http://210.75.249.4/handle/363003/1708
专题	中国科学院西北高原生物研究所
作者单位	1.Chinese Acad Sci, NW Inst Plateau Biol, Xining 810001, Peoples R China 2.Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China
推荐引用方式 GB/T 7714	Zhang, B. O.,Wang, Haiqing,Shen, Yuhu,et al. Isolation, analysis and prokaryotic expression of nitric oxide associated factor gene-NOA1 from potato (Solanum tuberosum L.)[J]. AFRICAN JOURNAL OF BIOTECHNOLOGY,2010,9(18):2606-2612.
APA	Zhang, B. O.,Wang, Haiqing,Shen, Yuhu,&Zhang, Huaigang.(2010).Isolation, analysis and prokaryotic expression of nitric oxide associated factor gene-NOA1 from potato (Solanum tuberosum L.).AFRICAN JOURNAL OF BIOTECHNOLOGY,9(18),2606-2612.
MLA	Zhang, B. O.,et al."Isolation, analysis and prokaryotic expression of nitric oxide associated factor gene-NOA1 from potato (Solanum tuberosum L.)".AFRICAN JOURNAL OF BIOTECHNOLOGY 9.18(2010):2606-2612.