摘要
为研究辣椒CaWRKY30转录因子与番茄斑萎病毒互作的响应机制,以辣椒湘研11为试验材料,通过RNA提取、RT-PCR、切胶纯化及克隆等试验获得CaWRKY30编码序列。生物信息学分析结果表明,CaWRKY30全长1122 bp,编码373个氨基酸,该基因编码的蛋白含有1个WRKY保守结构域和1个C_(2)H_(2)结构域,属于典型的Ⅱ(e)亚家族成员。系统发育分析表明,与马铃薯StWRKY22氨基酸序列的亲缘关系最近。本氏烟叶片亚细胞定位试验发现,CaWRKY30在本氏烟幼苗中定位于细胞核和细胞膜,并且导致细胞膜加厚。实时荧光定量PCR分析结果表明,观察番茄斑萎病毒机械摩擦接种辣椒后的病毒积累量,发现病毒积累量在接种1~14 d逐渐上升,在接种后14 d处于最大值,接种14 d后病毒积累量逐渐下降;同时,CaWRKY30受番茄斑萎病毒接种后诱导,在接种病毒1~14 d CaWRKY30表达量上调,14 d达到峰值,14 d以后表达量逐渐下降。综上,获得了CaWKRY30转录因子基因序列,该转录因子定位于细胞核和细胞膜,并初步阐述了辣椒CaWRKY30转录因子在番茄斑萎病毒胁迫下的表达趋势。
In order to study the response mechanism of pepper CaWRKY30 transcription factor and Tomato spotted wilt orthotospovirus,it was experimental materials with pepper Xiangyan 11.The CaWRKY30 coding sequence was obtained by RNA extraction,RT-PCR,split gel and cloning.Biological information analysis results showed that CaWRKY30 full length was 1122 bp,encoding 373 amino acids,the gene encoded protein contains 1 WRKY conservative domain and 1 C_(2)H_(2) domain,belonged to a typicalⅡ(e)subfamily member.System evolution analysis showed that the relative relationship with the potato StWRKY22 amino acid sequence was recently.It was found that CaWRKY30 was positioned in the nucleus and cell membranes in its cigarette seedlings,and leads to cell membranes.The results of Real-time fluorescence quantitative PCR analysis showed that the viral accumulation of Tomato spotted wilt orthotospovirus mechanical friction-vaccination was found that the viral accumulation was gradually increased from 1 to 14 days after inoculation,and virus accumulation reached its maximum in 14 days,after inoculation 14 days,viral accumulation gradually declined.At the same time,CaWRKY30 was induced by Tomato spotted wilt orthotospovirus,when the inoculation 1—14 days,the CaWRKY30 expression was raised,and the peak was reached in 14 days,the expression in 14 days gradually decreased.In summary,it obtained the CaWRKY30 transcription factor gene sequence,which was located in the nucleus and cell membrane,and preliminarily explained the expression trend of CaWRKY30 transcription factors under the stress of Tomato spotted wilt orthotospovirus.
作者
贾志强
许云玉
高雪
陶宏征
陈增敏
刘雅婷
李永忠
JIA Zhiqiang;XU Yunyu;GAO Xue;TAO Hongzheng;CHEN Zengmin;LIU Yating;LI Yongzhong(College of Plant Protection,Yunnan Agricultural University,Kunming 650201,China;College of Agronomy and Biotechnology,Yunnan Agricultural University,Kunming 650201,China;College of Life Science and Technology,Honghe University,Mengzi 661199,China;College of Tobacco Science,Yunnan Agricultural University,Kunming 650201,China)
出处
《华北农学报》
CSCD
北大核心
2022年第3期186-192,共7页
Acta Agriculturae Boreali-Sinica
基金
国家自然科学基金项目(31471828,31260451)
云南省重点项目(2018FA019)。
