遗传 ›› 2009, Vol. 31 ›› Issue (4): 426-433.doi: 10.3724/SP.J.1005.2009.00426

• 研究报告 • 上一篇    下一篇

盐胁迫下西伯利亚蓼茎叶正反消减文库的构建及初步分析

刘关君;刘明坤;许志茹;阎秀峰;魏志刚;杨传平   

  1. 东北林业大学林木遗传育种与生物技术教育部重点实验室, 哈尔滨 150040
  • 收稿日期:2008-07-18 修回日期:2008-09-04 出版日期:2009-04-10 发布日期:2009-04-10
  • 通讯作者: 杨传平

Construction and analysis of a forward and reverse subtractive cDNA library from leaves and stem of Polygonum sibiricum Laxm. under salt stress

LIU Guan-Jun;LIU Ming-Kun;XU Zhi-Ru;YAN Xiu-Feng;WEI Zhi-Gang;YANG Chuan-Ping
  

  1. The Laboratory of Forest Genetics and Breeding and Biotechnology of Ministry of Education, Northeast Forestry University, Harbin 150040, China
  • Received:2008-07-18 Revised:2008-09-04 Online:2009-04-10 Published:2009-04-10
  • Contact: MingKun Liu

摘要: 正向文库以3% NaHCO3胁迫48 h的材料(茎、叶)为实验方(tester), 以未胁迫的材料(茎、叶)为消减方(driver); 在负向文库中, 以未胁迫的材料(茎、叶)为实验方(tester), 以3% NaHCO3胁迫48 h的材料(茎、叶)为消减方(driver), 利用抑制性消减杂交技术(suppression subtractive hybridization, SSH)构建了3% NaHCO3胁迫下西伯利亚蓼茎叶正反消减文库。从文库中共获得2 282条有效EST序列, 其中从茎正向消减文库中获得598条, 从茎负向消减文库中获得490条, 从叶正向消减文库中获得627条, 从叶负向消减文库中获得567条。经BlastX序列比对后, 通过MIPs方法对EST功能进行分类并对茎叶正反消减文库做了比较, 其中茎与叶部除细胞救援与防御、转运组件中变化趋势相同外, 在代谢、能量、光合作用、蛋白合成、转录和信号传导等方面均表现为相反趋势, 另外通过荧光定量RT-PCR对12个潜在与盐胁迫相关基因进行定量分析, 结果显示12个基因在未经胁迫与盐胁迫处理后的西伯利亚蓼叶与茎部有着很大差异, 说明叶与茎部在NaHCO3条件下可能具有不同应答机制与分工, 这有助于进一步理解西伯利亚蓼分子耐盐机制。

关键词: 西伯利亚蓼, 消减文库, 差异表达基因, 盐胁迫

Abstract: Using cDNAs prepared from the leaves and stems of Polygonum sibiricum Laxm. treated with NaHCO3 stress for 48 h as testers and cDNAs from unstressed P. sibiricum leaves and stems as drivers library, suppression subtractive hy-bridization (SSH) was employed to construct a cDNA subtracted library, which contained 2 282 valid sequences including 598 ESTs in the stems forward SSH library and 490 ESTs in the stem reverse SSH library, 627 ESTs in the leaf forward SSH library and 567 in the leaf reverse SSH library. According to the functional catalogue of MIPs and the comparison of the reverse and forward SSH libraries of the stem and leaf, the responses to NaHCO3 stress were different between leaf and stem, except for the same trend in cell rescue defense and transport facilitation. The trend in the metabolism, energy, photo-synthesis, protein synthesis, transcription, and signal transduction was opposite. RT-PCR analysis demonstrated that the expression of 12 putative stress related genes in the NaHCO3-treated leaves and stems was different from that in the un-treated leaves and stems. This indicated that different mechanisms might be responsible for reactions of leaf and stem in P. sibiricum. The results from this study are useful in understanding the molecular mechanism of saline-alkali tolerance in P. sibiricum.