摘要
WRKY转录因子广泛参与植物对生物和非生物的胁迫应答反应,在植物防卫反应中起着重要作用。CaRKNIF1是我们从辣椒HDA149中分离得到的新WRKY转录因子基因(GenBank登陆号DQ180348),本研究通过RT-PCR扩增得到CaRKNIF1基因的开放阅读框,经由中间载体pEASY-T1 Simple,将其定向连接到表达载体pCHF3上,得到植物表达载体pCHF3-CaRKNIF1。采用电击法将pCHF3-CaRKNIF1导入农杆菌菌株LBA4404,农杆菌介导法获得了抗性番茄植株。PCR、Southern杂交和RT-PCR检测结果表明,CaRKNIF1基因已经成功整合到番茄基因组中,并正常表达。此研究为进一步通过CaRKNIF1基因提高转基因番茄的抗逆性奠定了基础。
WRKY transcription factors are extensivelyinvolved in plants responses to biotic/abiotic stresses,playing important roles in plant defense responses.CaRKNIF1 gene（GenBank No.DQ180348） is a new WRKY transcription factor isolated from pepper HDA149.In this study,we amplified the CaRKNIF1 ORF by RT-PCR using adaptor primers and cloned it into intermediate vector pEASY-T1 Simple.Then,we cut it from the intermediate vector and linked it to expression vector pCHF3 to form plant expression vector pCHF3-CaRKNIF1 which was later transferred into Agrobacterium tumefaciens LBA4404 by electroporation.In this way,kanamycin resistant tomato lines were obtained.PCR,Southern blot and RT-PCR analysis indicated that CaRKNIF1 gene had been successfully integrated into the genome of transgenic tomato lines and was expressed normally.This work laid good foundation for enhancing tomato resistance to different stresses by expression of the CaRKNIF1 gene.
出处
《分子植物育种》
CAS
CSCD
2010年第4期713-718,共6页
Molecular Plant Breeding
基金
国家重点基础研究发展计划(2009CB119000)
国家自然科学基金项目(30671412)
农业公益性行业科研专项(nyhyzx07-050)共同资助
关键词
WRKY转录因子
表达载体构建
番茄
遗传转化
WRKY transcription factor
Expression vector construction
Tomato
Genetic transformation
