干旱胁迫下黄檗幼苗 cDNA 消减文库的构建和分析.pdfVIP

生 物 工 程 学 报 Chin J Biotech 2008, February 25; 24(2): 198-202 Chinese Journal of Biotechnology ISSN 1000-3061 cjb@ © 2008 Institute of Microbiology, CAS CSM, All rights reserved 研究报告 cDNA 王慧梅, 王延兵, 祖元刚, 孙莲慧 , 150040 : 以干旱胁迫下的黄檗幼苗 cDNA 为 tester, 正常生长的黄檗幼苗 cDNA 为 driver, 利用抑制性消减杂交技术 (suppression subtractive hybridization, SSH)构建了干旱胁迫下黄檗幼苗的消减文库并对其进行了EST 序列分析。从消减 文库中随机挑取20 个阳性克隆, 提取质粒进行酶切和PCR 鉴定, 显示文库克隆的重组率大于95%, 插入片段大小大部 分集中在300~800bp 之间。随机挑取 816 个克隆进行测序, 得到265 个基因。将其进行同源性分析, 划分为 16 类。获 得了热激蛋白 70 、脱水响应蛋白(RD22) 、通用胁迫蛋白、金属硫蛋白(MTII), 晚期胚胎丰富蛋白(LEA14)等 44 种与干 旱胁迫相关的基因,它们涉及了植物的渗透调节、信号传递、转录调控、活性氧清除等方面。本研究为抗逆基因克隆和 系统研究干旱胁迫下黄檗基因的表达奠定了重要的理论基础。 : 干旱胁迫, 黄檗, 抑制性消减杂交, EST 序列分析 Construction and Analysis of Subtractive cDNA Library of Phellodendron amurense Under Drought Stress Huimei Wang, Yanbing Wang, Yuangang Zu, and Lianhui Sun Key Laboratory of Forestry Plant Ecology, Ministry of Education, Northeast Forestry University, Harbin 150040, China Abstract: With cDNA from Phellodendron amurense seedlings treated with drought stress as tester and cDNA from this plant in normal growth as driver, we construct cDNA subtracted library using suppression subtractive hybridization (SSH). In the library, the rate of recombination was 95%, the size of inserts was 300~800 bp. Two hundred and sixty-five new genes were obtained by DNA sequencing 816 positive clones picked randomly, and partitioned to 16 classes after nucleotide Blast and BlastX homological analysis against NT, NR, SWISSPROT, KEGG database. Forty-four drought stress associated genes, such as hea