番茄Sly-miR393基因超表达载体的构建及其靶基因鉴定
onstruction of Sly-miR393 Over-expression Vector and Verification of its Target Genes in Tomato
为了研究番茄Sly-miR393的功能，通过生物信息学方法从番茄基因组数据库中分析和预测Sly-miR393的前体序列及其潜在的靶基因，以基因组 DNA 为模板，克隆了番茄Sly-miR393 前体基因并将其亚克隆到pLP35s-100植物表达载体；同时，采用5'RACE技术验证Sly-miR393对预测靶基因mRNA的剪切作用及其剪切位点。结果表明，Sly-miR393的前体序列含有完整的茎环结构；成熟Sly-miR393与TIR1/AFB生长素受体同源基因之间具有识别位点。Sly-miR393在番茄体内对靶基因SlTIR1，SlTIR1-like1和SlAFB的转录本通过一个切割位点进行剪切降解；从而证实了生长素受体同源基因为Sly-miR393的靶基因。成功构建了以 CaMV35S 为启动子的植物表达载体pLP35S-pre-SlymiR393, 为深入研究Sly-miR393在番茄生长素信号转导中的功能奠定了基础。
o understand the function of Sly-miR393 in tomato, the precursor sequences and potential target genes of Sly-miR393 were identificated from tomato genome database by computational homology search method. The Sly-miR393 gene was amplified from the genomic DNA by PCR and cloned into plant expression vector pLP35s-100. Sly-miR393 guided-cleavage to putative target transcripts was validated using 5'RACE RT-PCR. In this study, our results indicated that the precursor sequence of Sly-miR393 contains the complete hairpin structure. TIR1/AFB auxin receptor genes contain recognition sites with high complementarities to Sly-miR393 sequence. In tomato, Sly-miR393 directs the cleavage of SlTIR1，SlTIR1-like1 and SlAFB mRNA, then auxin receptor homologous was validated to be as target of Sly-miR393. The pLP35s-pre-SlymiR393 vector containing Sly-miR393 gene was successfully constructed, which would provide significant evidence for further study of Sly-miR393 function in auxin signaling pathway in tomato.
李正国、林冬波、杨迎伍
分子生物学植物学遗传学
Sly-miR393靶基因生长素受体基因载体构建
Sly-miR393Target genesAuxin receptor geneConstruction
李正国,林冬波,杨迎伍.番茄Sly-miR393基因超表达载体的构建及其靶基因鉴定[EB/OL].(2012-05-09)[2025-08-02].http://www.paper.edu.cn/releasepaper/content/201205-129.点此复制
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