基于cDNA-AFLP发掘茶树被茶尺蠖取食诱导的相关差异基因及其表达特征

doi:10.3864/j.issn.0578-1752.2013.19.018

摘要/Abstract

摘要： 【目的】发掘茶树被茶尺蠖取食诱导的相关防御基因及其分子防御机制。【方法】利用cDNA-AFLP筛选茶树叶片被其主要害虫茶尺蠖取食后差异表达片段（transcript-derived fragment，TDF），并对其进行生物信息学分析和相关验证；运用qRT-PCR（quantitative real-time PCR）研究部分差异基因的表达规律。【结果】茶树被茶尺蠖取食后，共发现231条TDF，经PCR验证和测序后，成功获得134条EST（expressed sequence tags）序列，其中上调与下调表达的TDF数量分别为81和53条，各占测序获得序列的60.4%与39.6%；利用BLASTx在GenBank进行比对与功能分析可知，其中的23.1%与代谢相关，9.0%跟光合作用与能量相关，跟蛋白质合成与贮藏相关的序列各占8.2%，与信号转导相关的占6.0%，跟抗病与防御相关的占5.2%，但与细胞生长及细胞结构、转录因子和运输相关的数量很少，分别为1.5%、2.2%和3.0%；此外，还有6.7%的TDF功能不明确，有20.1%的无同源性序列；利用qRT-PCR（quantitative real-time PCR）对TDF 21F1、22D、27L3和28F分别进行了表达特征分析，结果表明这4个TDF代表的基因在茶树被茶尺蠖取食后均有不同程度诱导。【结论】通过cDNA-AFLP技术发掘了多个茶树被茶尺蠖取食诱导的差异表达基因片段，初步揭示了茶树被害虫取食而诱导的分子防御机制。

关键词: 茶树 , 茶尺蠖 , 取食诱导 , cDNA-AFLP , 防御机制

Abstract: 【Objective】 The objective of this study is to screen the defense genes in tea plant (Camellia sinensis L.) induced by tea looper (Ectropis oblique).【Method】Tea leaves fed by the looper were analyzed by cDNA-AFLP(cDNA-amplified fragment length polymorphism). Transcript-derived fragments (TDF) were screened and confirmed. After that, bioinformatics analysis was followed. And the expression patterns of the differential genes were performed by qRT-PCR.【Result】The results showed that 134 EST(expressed sequence tags) were obtained by sequencing and identifying from the selected 231 TDFs. Among them, 81 TDFs were up-regulated, and 53 were down-regulated, which accounted for 60.4% and 39.6% of the obtained fragments, respectively. According to BLASTx result of TDFs, a large group (23.1%) of these genes shared high homology with genes involved in metabolism, 9.0% genes with functions related to photosynthesis and energy, 8.2% genes with functions related to protein synthesis and storage, signal transduction, 5.2% genes with functions related to disease/defense. But only few genes involved in cell growth and structure, transcription factor and transporters and the ratios were 1.5%, 2.2%, and 3.0%, respectively. Moreover, sequences with unknown proteins occupied 6.7%; and 20.1% sequences with no similarity against the database. TDF-21F1, TDF-22D, TDF-27L3 and TDF-28F were confirmed by qRT-PCR analysis, further result indicated that the expression of the defense genes responsive to Ectropis oblique play significant regulation roles in adapting to negative environment for tea plant.【Conclusion】Using cDNA-AFLP method to analyze Ectropis oblique feeding tea plant and get the TDFs, it could supply valuable information to the study of the molecular mechanism between tea and pest.

Key words: Camellia sinensis , Ectropis oblique , feeding-induced , cDNA-AFLP , defense mechanism

曹士先, 程曦, 蒋正中, 盛亮, 上官明珠, 邓威威, 韦朝领. 基于cDNA-AFLP发掘茶树被茶尺蠖取食诱导的相关差异基因及其表达特征[J]. 中国农业科学, 2013, 46(19): 4119-4130.

CAO Shi-Xian, CHENG Xi, JIANG Zheng-Zhong, SHENG Liang, SHANG Guan-Ming-Zhu, DENG Wei-Wei, WEI Chao-Ling. Differential Genes Expression in Tea Plant (Cameilla sinensis L.) Induced by Ectropis oblique Feeding Based on cDNA-AFLP[J]. Scientia Agricultura Sinica, 2013, 46(19): 4119-4130.

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