烟草野火病菌hrpZ基因的克隆及蛋白的原核表达

Cloning and Expression of hrpZ Gene from Pseudomonas syringae pv. tabaci

采用PCR方法从烟草野火病病原菌Psta218中扩增harpin编码基因hrpZ,将其克隆到原核表达载体pGEX-4T-1上,获得重组表达质粒pGEX-hrpZPsta。将重组质粒pGEX-hrpZPsta转化至大肠杆菌BL21(DE3)菌株,得到重组大肠杆菌BL21/pGEX-hrpZPsta。经IPTG诱导得到14.7 kD的蛋白质。该蛋白质与其他已发现的harpins一样,能够使烟草等植物产生过敏性坏死反应、富含甘氨酸、热稳定以及对蛋白酶K敏感。

The hrpZ gene was amplified from genome of Pseudomonas syringae pv.tabaci Psta218 by PCR and then constructed expression vector pGEX-hrpZPsta with regular molecular cloning operation.The recombinant plasmid was transformed into BL21（DE3）.Recombinant protein was induced by Isopropylthio-β-D-Galacgoside（IPTG）.The molecular mass of the fusion protein is 14.7 kD analyzed by SDS-PAGE.The protein,similar to the other known harpins,was heat-stable,which contained abundant glycine（G）,but had no cysteine.Furthermore,this protein was sensitive to protease K and able to trigger hypersensitive response（HR） in common tobacco.