【摘要】 电子克隆是基因克隆的新策略。以小麦胞质葡萄糖 6 磷酸脱氢酶cDNA (Tagpdl克隆 )序列为信息探针 ,在GenBank水稻nr数据库中找到高度同源的水稻基因组序列 ,通过人工序列拼接及RT PCR确认得到了水稻该基因的全长cDNA序列 ,命名为OsG6PDH。OsG6PDH与小麦Tagpdl克隆的DNA一致率为 88% ,推导的氨基酸序列与小麦、番茄、烟草的胞质葡萄糖 6 磷酸脱氢酶基因的一致率分别为 89%、79%、80 %。经RT PCR表达谱分析 ,OsG6PDH在水稻幼穗、胚、根、叶中都有表达 ,在幼穗与根中表达略高。另外 ,讨论了利用水稻基因组信息的电子克隆方法克隆水稻功能基因的可行性更多还原

【Abstract】 In silico cloning was a new strategy of gene cloning developed with the development of genome, EST projects and bioinformatics. Using wheat glucose 6 phosphate dehydrogenase cDNA (clone:Tagpdl) sequence as a querying probe, one highly homologous BAC clone sequence was obtained from rice sequence database of GenBank and the putative cDNA sequence of rice glucose 6 phosphate dehydrogenase was assembled according to the wheat clone. Futhermore, the full length cDNA of rice glucose 6 phosphate dehydrogenase was cloned by RT PCR with two primers designed based on this assembled cDNA sequence. Since this fragment contained a complete ORF of 1515bp with a stop codon in its upstream and poly(A) signal in its downsteam, it could be concluded that a full length gene (GenBank accession number AY078072), which was named as OsG6PDH . Homology analysis of OsG6PDH showed a 88% identity with wheat and the deduced amino acid showed 89%, 79% and 80% omology with G6PDH from wheat, tomato and tobacco respectively. OsG6PDH was expressed in inflorescence, embryo, root and leaf of rice, with a slightly higher in inflorescence and root. It was also discussed in this paper that the application of in silico cloning in the isolation of functional genes from rice.更多还原