水稻新基因OsRwp34在大肠杆菌中表达的毒害作用

OsRwp34,a New Rice Gene Which Expresses a Toxic Product to E.coli Growth

在水稻高温诱导cDNA文库中克隆了一个新的水稻基因O sRwp34,同源性分析表明O sRwp34是一典型的孤儿基因。为了研究O sRwp34的功能,构建了O sRwp34的表达载体pET-28(a)OsRwp34,并转化大肠杆菌菌株BL21(DE3),用IPTG诱导并定时从培养液中取样以检测OD600值和用平板记数法测定活菌数,结果发现在诱导30 m in后宿主菌开始大量死亡,表明O sRwp34的表达产物使宿主菌死亡。随后构建了1个N末端缺失15个氨基酸残基和2个C末端分别缺失12和47个氨基酸残基的O sRwp34缺失体,IPTG诱导后都没有出现毒性作用,推测N和C末端氨基酸残基的同时存在是OsRwp34的毒性作用所必须的。其详细机理有待进一步研究。

OsRwp34 was isolated in a cDNA library which was constructed from rice （ Oryza sativa） seedling under high temperature（45℃ ） stress. In order to study the function of OsRwp34 ,we constructed an expression vector pET-28 （a）OsRwp34 which included OsRwp34 gene and transformed it into the E. coli strain BL21 （DE3）. The bacteria was incubated in LB medium and induced by IPTG,and the host bacteria was found to be dying rapidly in 30 min through the OD600 monitor and quantitative plating, The resuit suggested that the expression product of OsRwp34 caused the death of the bacteria. Continuously, three deletion mutants,one of them deleted 15 aa of N terminate,the other two deleted 12 aa and 47 aa of C terminate respectively ,were constructed. But the toxic effect to the bacteria growth was not found after the induction of IPTG. It indicates that the exist of both N and C terminates are necessary for the toxic effect of OsRwp34. A further study will be required on the toxic mechanism of OsRwp34.