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浙江大学学报（农业与生命科学版）
浙江大学学报(农业与生命科学版)  2015, Vol. 41 Issue (2): 153-159    DOI: 10.3785/j.issn.1008-9209.2014.05.201
生物科学与技术     
青花菜转录因子基因BoWRKY2的克隆与表达分析
蒋明*， 陈贝贝， 管铭， 李金枝， 黄笑梅， 顾云吉
台州学院生命科学学院，植物进化生态学与保护省重点实验室/生态学省重点学科,浙江 椒江 318000
Cloning and expression analysis of a transcription factor gene BoWRKY2 from broccoli.
Jiang Ming*, Chen Beibei, Guan Ming, Li Jinzhi, Huang Xiaomei, Gu Yunji
(Zhejiang Provincial Key Laboratory of Plant Evolutionary Ecology and Conservation/Ecology Key Discipline of Zhejiang Province, College of Life Sciences, Taizhou University, Jiaojiang 318000, Zhejiang, China)
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摘要： 以青花菜为材料，克隆到1个WRKY转录因子基因BoWRKY2，在序列分析的基础上，利用反转录聚合酶链反应（reverse transcription-polymerase chain reaction, RT-PCR）检测其在核盘菌和霜霉菌侵染下的表达模式。结果表明：BoWRKY2的基因组全长为1 507 bp，具有2个内含子，编码区全长为987 bp；BoWRKY2编码328个氨基酸，具有1个WRKYGQK残基和C-X5-C-X23-H-X1-H锌指结构，WRKY结构域与甘蓝型油菜的最为相似，仅存在1个氨基酸残基的差异。进化分析结果表明，BoWRKY2与同为十字花科的甘蓝型油菜、拟南芥、深山南芥、荠菜和盐芥聚为一组，支持率达97%。RT-PCR结果表明，BoWRKY2的表达受霜霉菌和核盘菌的诱导，二者的表达模式相似，在6 h和12 h时表达量增加，但在24 h后下降，暗示BoWRKY2与2种病菌的早期抗性反应相关.
Abstract: Broccoli (Brassica oleracea var. italica), which belongs to Cruciferae family, is a cash crop widely cultivated in China, and it is regarded as one of the most consumed vegetables in the world. The flower head of broccoli is rich in minerals, vitamins, fibers as well as anti-oxidants, so it is recognized as a healthy vegetable with anti-cancer properties. As a major broccoli production center in China, the average plantation areas reach to 8 000 hm2 in Taizhou of Zhejiang Province. However, broccoli cultivation suffered from plant diseases of downy mildew and stalk break which were caused by Hyaloperonospora parasitica and Sclerotinia sclerotiorum, respectively, resulting in yield and quality loss. WRKY transcription factors played important roles in plant stress responses, and WRKY domains were defined as an approximately 60-amino acid motif named WRKYGQK as well as a zinc finger structure of C-X4-5-C-X2223-H-X1-H at their C-terminus. The WRKY domain binds specifically to DNA sequence of (T)(T)TGAC(C/T) known as the W-box which exists in defense-related promoters. Enhanced disease resistance by overexpression of WRKY genes in different crop plants has been reported in recent years. In this study, a WRKY gene designated as BoWRKY2, was isolated from B. oleracea var. italica. Based on sequence analysis, expression patterns of BoWRKY2 were detected using reverse transcription-polymerase chain reaction (RT-PCR) method while challenged by H. parasitica and S. sclerotiorum, respectively. The results showed that the genome DNA sequence was 1 507 bp in length with two introns and a complete coding sequence of 987 bp, and the length of two introns were 425 and 95 bp, respectively; BoWRKY2 encoded 328 amino acids with a WRKYGQK residue and a zinc finger structure of C-X5-C-X23-H-X1-H. The WRKY domain located between 247 and 313 residues, and several DNA binding sites were found at sites of 66, 67, 69, 71, 74, 281, 282, 283 and 317. The WRKY domain was similar to that of oilseed rape with only one amino acid residue difference between them. Phylogenetic analysis indicated the BoWRKY2 was grouped with other Cruciferae plants such as B. napus, Arabidopsis thaliana, A. lyrata, Capsella rubella and Eutrema salsugineum, with 97% confidence. RT-PCR results revealed that the BoWRKY2 was induced by both H. parasitica and S. sclerotiorum with similar expression patterns. The expression level both increased at 6 h and 12 h, and decreased after 24 h, indicating the resistance responses of BoWRKY2 against two plant fungi. In a word, the cloning and expression analysis of BoWRKY2 gene lay the foundation for further studies in gene function identification and molecular breeding of broccoli.
出版日期: 2015-03-20
CLC:  Q 78  
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蒋明
陈贝贝
管铭
李金枝
黄笑梅
顾云吉

引用本文:

蒋明，陈贝贝，管铭，李金枝，黄笑梅，顾云吉. 青花菜转录因子基因BoWRKY2的克隆与表达分析[J]. 浙江大学学报(农业与生命科学版), 2015, 41(2): 153-159.

Jiang Ming, Chen Beibei, Guan Ming, Li Jinzhi, Huang Xiaomei, Gu Yunji . Cloning and expression analysis of a transcription factor gene BoWRKY2 from broccoli.. Journal of Zhejiang University (Agriculture and Life Sciences), 2015, 41(2): 153-159.

链接本文:

http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2014.05.201    或    http://www.zjujournals.com/agr/CN/Y2015/V41/I2/153

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