萝卜抗真菌蛋白基因Rs-AFPs在大肠杆菌中的表达及其转化番茄的研究

The Expression of Raphnus sativus-Antifungal Protein Gene and Its Transformation into Tomato

通过基因工程手段将萝卜抗真菌蛋白基因Rs AFP1( 2 ) 插入大肠杆菌表达载体pTrxFus中并诱导表达 ,其融合表达产物约 2 2kD ,约占总可溶蛋白的 0 .4 5 % (0 .5 % )。抑菌活性试验表明 :重组Rs AFP1( 2 ) 有抑菌活性 ,其中Rs AFP2 较Rs AFP1抑菌活性强。构建了Rs AFP2 基因的植物表达载体pBIAFP2 。利用农杆菌介导法将pBIAFP2 导入番茄中 ,并诱导出完整的植株 32株。对其中 2 8株进行PCR和PCR SouthernBlot检测 ,其中 17株呈阳性。对经PCR SouthernBlot检测呈阳性的植株进行SouthernBlot检测 ,6株呈阳性。

c cysteine rich,antifungal proteins.The Rs AFP 1(2) genes were inserted into the vector pTrxFus and transferred into E.coli GI724.The fusion protein from recombinant pTrxAFP 1(2) was about 22kD and taking up 0.4 and 0.5 percent of the total proteins respectively.Inhibition experiment showed that the expressed products had anti fungal activity to Colletotrichum musae.Expressed products from pTrxAFP 2 showed much stronger inhibitory activity than that from pTrxAFP 1.A plant expression vector pBIAFP 2 was constructed, which had been transferred into tomato by Agrobactrium mediated transformation.32 plantlets had been obtained and 28 of them detected by PCR and PCR Southern Blot.The results showed that 17 of them were positive,and the results of Southern Blot showed 6 of them were positive.