砀山酥梨萼片发育的ssh文库构建及est序列分析-安徽农业大学.pdfVIP

安徽农业大学学报, 2013, 40(3): 454-459 Journal of Anhui Agricultural University 网络出版时间：2013-4-26 15:37:40 [URL] /kcms/detail/34.1162.S1537.029.html 砀山酥梨萼片发育的 SSH 文库构建及 EST 序列分析 贾 胜，贾 兵，朱立武∗ (安徽农业大学果树学重点实验室，合肥 230036) 摘 要：为了寻找梨幼果期萼片发育中差异表达基因，为梨幼果萼片脱落分子机制研究提供依据，以砀山酥梨 萼片脱落开始期幼果为材料，利用抑制性消减杂交（suppression subtractive hybridization, SSH ）技术构建砀山酥梨 幼果期宿萼果与脱萼果差异表达cDNA 文库。随机挑取200 个阳性克隆，经菌液PCR 检测及测序后得到 185 个EST 序列片段， 其中小于100 bp、大于300 bp 以及介于两者之间的EST 片段分别占3%、96%和1%。将这些片段在 NCBI 上进行BLAST 比对，得到调节因子（regulator ）、蛋白质（protein ）和相关酶类（related enzyme ）3 类代谢 途径相关物质，分别占 21% 、63%和 16%。其中，在 Gene Ontology 网站上比对到 3-异丙基苹果酸脱氢酶 （3-isopropylmalate dehydrogenase ）和N- 酰基-L-氨基酸氨基水解酶（N-acyl-L-amino acid amidohydrolae ）2 种酶， 推测可能是砀山酥梨幼果期萼片发育代谢途径中的重要的相关酶。 关键词：砀山酥梨；萼片；抑制差减杂交技术；SSH 文库；序列分析 中图分类号：S661.2 文献标识码：A 文章编号：1672−352X (2013)03−0454−06 SSH library construction and EST sequence analysis on the development of the calyx of ‘Dangshansu’ pear JIA Sheng, JIA Bing, ZHU Li-wu (Key Laboratory of Pomology, Anhui Agricultural University, Hefei 230036) Abstract: The aim of this experiment is to investigate the differentially expressed genes on the development of calyx in the fruitlet period of pear and to provide evidence for the research of molecular mechanisms of its ca- lyx shedding. A subtracted cDNA library about the different expressions between the calyx-setting and the ca- lyx-shedding fruitlet was constructed by using suppression subtractive hybridization(SSH), and the fruitlets of ‘Dangshansu’ pear at the time of calyx shedding were used as materials. A total 200 positive clones were randomly screened out, and 185 expressed se