寄生虫与医学昆虫学报2012，Vol.19Issue(4)：228-234,7.DOI:10.3969/j.issn.1005-0507.2012.04.006
家蝇抗菌肽defensin基因的克隆、原核表达纯化及抑菌、抑肿瘤活性鉴定
CLONING, PROKARYOTIC EXPRESSION AND PURIFICATION OF DEFENSIN GENE OF MUSCA DOMESTICA ANTIBACTERIAL PEPTIDE, AND IDENTIFICATION OF BACTERIOSTATIC AND CARCINOSTATIC ACTIVITIES THEREOF
摘要
Abstract
The genie sequence of defensin gene ( GenBank No. AY260152) was used to design specific primers for cloning mature defensin (MD). Total RNA was extracted from Musca domestica larva and MD was then amplified by PCR. PCR product was cut with Bam HI and Xhol and put into pGEX-6P-l vector. Recombinant expression plasmid pGEX-6P-l/ MD was constructed and target fragment was put into pGEX-6P-l vector stably through double digestion. The resultant recombinant plasmid (pGEX-6P-l/MD) was transformed into BL21 (DE3) . The fusion protein was expressed after IPTG induction at 25癈 , and purified by GST-Sefinose affinity chromatography. SDS-PAGE results showed that recombinant defensin was produced after IPTG induction. Antibacterial experiments indicated that the fusion recombinant protein had certain inhibitory effect on the growth of E. coli on tumor cells of K562 at the first time. Plasmid pGEX-6P-l/MD protein was cloned and expressed successfully, which testified the protein had inhibitory and damaging effect on K562 tumor cells. All that aforesaid provided a basis for the researches of antibacterial peptide drugs from M. domestica defensin.关键词
Defensin/抗菌肽/基因克隆/原核表达/亲和层析/肿瘤细胞Key words
Defensin/ Antibacterial peptide/ Gene clone/ Prokaryotic expression/ Affinity chromatography/ Tumour cells引用本文复制引用
焦莉萍,杨敏,侯养全,赵瑞君,原发家,杜斌,王文建..家蝇抗菌肽defensin基因的克隆、原核表达纯化及抑菌、抑肿瘤活性鉴定[J].寄生虫与医学昆虫学报,2012,19(4):228-234,7.基金项目
山西省高校科技研究开发项目(200613011) （200613011）
山西省科技攻关项目(2006031087-04) （2006031087-04）
太原市科学技术发展计划项目(081049) （081049）
