捻转血矛线虫ABC转运蛋白的基因克隆、原核表达及纯化

doi:10.3969/j.issn.1674-7968.2020.10.011

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Abstract Haemonchus contortus is one of the most important parasites of ruminants such as Ovis aries and Capra hircus, and can bring huge economic losses to the breeding industry. In preliminary transcriptome study of present research group, the resistant gene ATP-binding cassette (ABC) transporter gene (GenBank No. MT478136) was identified from the resistant strain of Haemonchus contortus. The purpose of this study was to explore the bioinformatics character of the ABC transporter gene, to establish a system of soluble expression and purification for ABC transporter, and to lay the foundation for further exploration of ABC transporter-involved drug resistance mechanism. The online software was used to predict the protein structure and antigen epitope. The results showed that the molecular formula of ABC was C₉₂₈H₁₅₀₂N₂₇₀O₂₆₅S₇, which was a hydrophilic protein without transmembrane region and signal peptide. The secondary structure was mainly extended chains. The prediction of antigenic epitopes indicated that ABC transporter had many antigenic epitopes with high antigenicity. The ABC transporter gene was amplified by reverse transcription PCR (RT-PCR), and the recombinant plasmid pET30a(+)-ABC was constructed by restriction digestion. The plasmid was transformed into Escherichia coli BL21 to optimize inducing conditions. It was found that 1 mg/mL recombinant ABC transporter could be expressed with 0.6 mmol/L isopropyl β-D-thiogalactoside (IPTG) at 37 ℃ for 7 h. In this study, recombinant ABC transporter was successfully expressed and purified in Haemonchus contortus, which could provide a reference for further mechanism study on potential drug resistance and immunogenicity.

Key words： Haemonchus contortus ATP-binding cassette (ABC) transporter gene Bioinformatics analysis Prokaryotic expression Protein purification

Received: 11 March 2020

ZTFLH:

S855.9

Corresponding Authors: * wwl.imau@163.com; lcx.imau@163.com

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	Articles by authors
	ZHAO Xue-Liang
	WANG Shu-Yi
	SUN Ke
	Huhe Bateer
	WANG Wen-Long
	LIU Chun-Xia

Cite this article:

ZHAO Xue-Liang,WANG Shu-Yi,SUN Ke, et al. Gene Cloning, Prokaryotic Expression and Purification of ABC Transporter in Haemonchus contortus[J]. 农业生物技术学报, 2020, 28(10): 1830-1836.

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http://journal05.magtech.org.cn/Jwk_ny/EN/10.3969/j.issn.1674-7968.2020.10.011 OR http://journal05.magtech.org.cn/Jwk_ny/EN/Y2020/V28/I10/1830

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