Expression, purification and antibody preparation of flagellin FlaA from Vibrio alginolyticus strain HY9901

	Expression, purification and antibody preparation of flagellin FlaA from Vibrio alginolyticus strain HY9901
	Xia, LQ; Jian, JC; Wu, ZH; wuzh@gdou.edu.cn; jianjc@gmail.com; Liang, HY; Lu, YS
刊名	LETTERS IN APPLIED MICROBIOLOGY
	2010
卷号	50 期号:2 页码:181-186
关键词	Antibody Preparation Expression Flagellin Flaa Gene Immunogold Electron Microscopy Purification Vibrio Alginolyticus
ISSN号	0266-8254
英文摘要	Aims: The main aims of this study were to clone and express flagellin flaA gene from Vibrio alginolyticus strain HY9901, also to prepare mouse anti-FlaA polyclonal antibody for future pathogen or vaccine study. Methods and Results: The full-length flaA gene was amplified by PCR with designed primers. The open reading frame of flaA gene contains 1131 bp, and its putative protein consists of 376 amino acid residues. Alignment analysis indicated that the FlaA protein was highly conserved. SDS-PAGE indicated that the FlaA protein was successfully expressed in Escherichia coli BL21 (DE3). Then, the recombinant FlaA protein was purified by affinity chromatography, and the mouse anti-FlaA serum was produced. The expression of flaA gene was verified by various immunological methods, including western blotting, enzyme-linked immunosorbent assay (ELISA) and immunogold electron microscopy (IEM). Conclusions: Flagellin flaA gene was cloned and identified from V. alginolyticus HY9901, the recombinant FlaA protein was expressed and purified, and high-titre FlaA protein-specific antibody was produced. Western blot analysis revealed that the prepared antiserum not only specifically react to FlaA fusion protein, but also to natural FlaA protein of V. alginolyticus. The expressed FlaA protein was demonstrated, for the first time, as the component of flagella from V. alginolyticus by IEM. Significance and Impact of the Study: This study may offer important insights into the pathogenesis of V. alginolyticus, provide a base for further studies on the diagnosis and evaluation that whether the FlaA protein could be used as an effective vaccine candidate against infection by V. alginolyticus and other Vibrio species. Additionally, the purified FlaA protein and polyclonal antibody can be used for further functional and structural studies.
学科主题	Biotechnology & Applied Microbiology ; Microbiology
URL标识	查看原文
资助项目	海洋生物
WOS记录号	WOS:000273316800008
公开日期	2011-07-03
内容类型	期刊论文
源URL	[http://ir.scsio.ac.cn/handle/344004/5442]
专题	南海海洋研究所_海洋生物
通讯作者	wuzh@gdou.edu.cn; jianjc@gmail.com
推荐引用方式 GB/T 7714	Xia, LQ,Jian, JC,Wu, ZH,et al. Expression, purification and antibody preparation of flagellin FlaA from Vibrio alginolyticus strain HY9901[J]. LETTERS IN APPLIED MICROBIOLOGY,2010,50(2):181-186.
APA	Xia, LQ.,Jian, JC.,Wu, ZH.,wuzh@gdou.edu.cn.,jianjc@gmail.com.,...&Lu, YS.(2010).Expression, purification and antibody preparation of flagellin FlaA from Vibrio alginolyticus strain HY9901.LETTERS IN APPLIED MICROBIOLOGY,50(2),181-186.
MLA	Xia, LQ,et al."Expression, purification and antibody preparation of flagellin FlaA from Vibrio alginolyticus strain HY9901".LETTERS IN APPLIED MICROBIOLOGY 50.2(2010):181-186.