花生种仁特异表达基因的初步筛选.pdfVIP

第 29 卷第4 期 江西农业大学学报 Vol. 29 ,No. 4 2007 年 8 月 Acta Agrículturae Universitatìs Jìangχjensjs Aug. ,2007 文章编号:1000 -2286(2∞7)04 -0680 -05 花生种仁特异表达墓园的初步筛选 蔡宁波，赵永莉，潘建菁，黄湘文，张君诚，庄伟建· {福建农栋大学油料作毒草研究所，福建福州 350自由2) 摘要:应m抑刽消减杂交技术，以花生矜仁cDNA 作为消减杂交约试验方〈如ster) , ~花生果皮 cDNA 作为驱动 方(driver) 进行消毒草杂交a 经过两轮拇牵U PCR 后，将第 Z 吹 PC丑产物与 pGEM-TEa町载体梧连，电击转化大 扇挎富，成功构建李宁仁特异表达 cDNA 文库，所长出菌落中 9 1. 2% 为白色克隆，采用 PCR 法对阳佐克运进行 鉴定，发王军其中单一扩增条警剖克娃约占 86.5% ，片段大小集中分布于2∞ -8阔地之降。窍花生种仁。D;II A 探针和果1t cDNA 靠在针分部对文库高密度杂交点阵膜进行1R1每 Northern 杂交检测。根据虽是交结果，如步从文 库中挑滚出 254 于差异点思花生种仨侍弄表达基因的相步声事选，为了解花生产量、品罢形成梧关始重要功能基 因，也为将来应用撞物圣因工程手段改良花生产量和品质奠定基础。 关键词.花生z 种仁:消接杂交:基因筛选 牢固分类号:Q781 ;5565. 2 文献标识到达 Primary Screening of DifferentialIy Expressed Genes in the Kernel of Peanut CAI Ning - bo ,ZHAO Yong -li. ,PAN Jian - jing , HUANG Xiang - wen ,ZHANG Jun - chen ,ZHUANG Wei - jian (Institute of Oil Crop , FAFU , Fuzhou 350002 , C毡皿a) Abstract: To screen the specífic genes în 世，. k.rneI of p臼R时， a differentiaUy 臼P野ssed cDNA lihrary was constructed by usin吕 suppression subtractive hyhridizatìon (SS日).For.uh桂Rct臼e hybridization , cDNA fro盟 the kernel of pe回国 W酷 used as the tester t 四d cDNA fro耻 the pod of peaDut was used as 也e drìver~ Af ter two tímes of subtractive hyl立idization and two times of nested PC且，也e prodUCIS of the second PCR ampli fication were Ìnserted 国to pGEM - T Easy vectors to transform the E. coli , a differentially expressed cDNA li brary of peanut kernel was suc臼ssfully construc恒d. 91.2% of the colonies w町e white τhe positive recombi 11811坦 were confirmed 也y P巳R ， 86.5%