质粒营救法和TAIL-PCR法获得水稻 T-DNA旁邻序列的效率比较.pdfVIP

农业生物技术学报 Journal of Agricultural Biotechnology 2004,12 1冤院 13~ 18 ·研究论文· 质粒营救法和TAIL-PCR 法获得水稻T-DNA 旁邻序列的效率比较 帆 金维正 陈双燕 吴运荣 刘非燕 吴 平* （浙江大学生命科学院，植物生理学与生物化学国家重点实验室，杭州310029 ） 摘要：获得突变体中T-DNA 的旁邻序列是通过反向遗传学进行功能基因研究的重要手段。实验比较了用质粒营救法和 TAIL-PCR 法获得T-DNA 在水稻( )基因组中旁邻序列的效率。利用质粒营救法获得T-DNA 在水稻中的旁邻序列 效率可高达90%左右，而利用TAIL-PCR 法的效率只有62% 。质粒营救法和TAIL-PCR 法获得的旁邻序列中水稻基因组DNA 的效率分别是80%左右和89%左右。根据实验结果，质粒营救法从转基因植株获得的旁邻序列，水稻基因组DNA 的效率是 72% ，比TAIL-PCR 法 18%。 ： ； ； 关键词 水稻 T-DNA ；质粒营救法 TAIL-PCR Comparison of the Efficiency Between Plasmid Rescue and TAIL-PCR Methods for Obtaining Flanking Sequence of T-DNA Tag Insertion in Rice Genome ZHANG Fan JIN Wei-Zheng CHEN Shuang-Yan WU Yun-Rong LIU Fei-Yan WU Ping* (State Key Laboratory of Plant Physiology and Biochemistry, College of Life Sciences, Zhej iang University, Hangzhou 310029, China) To obtain flanking sequence of T-DNA tag insertion in genome is a strategy for investigating functional genomics by using reverse genetics. In this study the plasmid rescue and tail-PCR methods were compared for their efficiency to obtain the flanking sequences T-DNA tag insertion in rice ( ) genome. The results from T-DNA insertion lines showed that the efficiency of plasmid rescue was about 90% , and that of TAIL-PCR was 62% . About 80% of flanking sequence of T-DNA tag insertion in rice genome by plasmid rescue was rice genome, while about 89% by TAIL-PCR. Therefore