小麦Wcor719基因的原核表达及转化烟草和棉花提高抗冷性研究  

PROKARYOTIC EXPRESSION OF Wcor719 GENE FROM WHEAT AND FREEZING TOLERANCE OF TRANSGENIC TOBACCO AND TRANSGENIC COTTON

作　　者：李晓荣[1] 王冬梅[1] 李建平[1] 李静[1] 张帅[1] 陈国华[1] 黄乐平[1] 

机构地区：[1]新疆农作物生物技术重点实验室/新疆农业科学院核技术生物技术研究所,新疆乌鲁木齐830091

出　　处：《核农学报》2012年第3期420-426,共7页Journal of Nuclear Agricultural Sciences

基　　金：新疆维吾尔自治区重大专项(200731138-3);新疆农作物生物技术重点实验室开放课题(XJYS0302-2011-04)

摘　　要：为研究Wcor719基因的抗寒功能,获得具有抗寒性的棉花新种质,构建小麦冷诱导基因原核表达载体Wcor719-pET28a进行蛋白表达,SDS-PAGE分析表明其诱导蛋白大小约为24.0kD,表达量在诱导4h时达到最大,且与IPTG诱导浓度无明显相关性。同时通过农杆菌介导的叶盘法将构建的Wcor719-pCambia1301植物表达载体转入烟草(Nicotiana tabacum)SR-1中,T1代转基因烟草植株经过抗生素筛选和PCR检测后进行低温胁迫,测定抗寒生理指标,结果显示转基因植株的脯氨酸和可溶性糖含量均比对照有所提高。该基因经过功能验证后用花粉管通道法转入棉花(Gossypium hirsutum)新陆中49,T0代、T1代经过田间检测及PCR检测后研究T2代转基因棉花低温胁迫下的抗寒性,结果显示,转基因棉花脯氨酸含量和可溶性糖含量均在第5天达到峰值,且明显高于对照;而丙二醛含量低于对照,初步证明转基因烟草和棉花的抗寒性均有所提高,由此表明通过转冷调节基因来提高植物的抗寒性具有良好的应用前景。A prokaryotic expression vector of Wcor719-pET28a from wheat was constructed to study the freezing tolerance function of Wcor719,and obtain new cotton germplasm tolerant to freezing.The SDS-PAGE analysis showed that the fusion protein was 24.0kD,and the highest expression amount was induced at 4h,moreover it had no obvious relevance with IPTG concentration.We transferred the Wcor719-1301 plant expression vector to tobacco（Nicotiana tabacum） SR-1 by Agrobacterium-mediated method,and then use antibiotic selection and PCR detection to obtain transgenic tobacco.Detecting of the T1 transgenic tobacco leaves showed that the proline content and soluble sugar of transgenic tobacco were higher than those of wild tobacco at low temperature treatment.Then the Wcor719-2301 was transferred to cotton（Gossypium hirsutum） by the pollen tube pathway,and the low temperature tolerance of T2 transgenic cotton was studied at 4℃.The result showed that the proline content and soluble sugar content of transgenic cotton reached peak value at the 5th days after treatment,which was higher than those of non-transgenic cotton.However,the MDA content of transgenic cotton was less than that of non-transgenic cotton,which proved that the freezing tolerance in the transgenic plants had been increased.It is indicated that it was a good method to increase the freezing tolerance by transferring COR genes.

关 键 词：Wcor719基因 蛋白表达 转基因烟草 转基因棉花 抗寒性 

分 类 号：S572[农业科学—烟草工业] S562[农业科学—作物学] S512.1