论文部分内容阅读
microRNA(miRNA)是生物中一类起负调控作用的非编码的小分子RNA,主要在转录后水平上调节生物体的生长与发育。直接克隆法是鉴定植物中miRNA最常用也是最有效的方法之一。利用直接克隆法分离了陆地棉开花后0~10 d(days post anthesis,DPA)胚珠中总RNA,从中筛选17~24 nt小分子RNA构建cDNA文库,对初筛得到的阳性克隆进行测序。通过与miRNA数据库比对,最终获得4个保守的miRNA。进一步对他们的转录表达进行分析,发现所有miRNA在棉花胚珠纤维不同发育时期均能表达。其中miR167a、miR172c和miR394b在棉花幼苗的根茎叶中有不同程度的表达。预测得到55个miRNA的靶基因,多数靶基因编码转录因子、代谢酶类以及发育相关蛋白,进一步证明miRNA参与调控棉花纤维的形态建成、发育等各项生理活动。
MicroRNAs (miRNAs) are a class of non-coding small RNAs that play a negative regulatory role in the organism and mainly regulate the growth and development of the organism at post-transcriptional level. Direct cloning is one of the most commonly used and most effective methods for identifying miRNAs in plants. The direct total RNA was isolated from ovules 0 to 10 days (days post anthesis) of Upland cotton by direct cloning method. A 17-24 nt small RNA was screened to construct a cDNA library. The positive clones were sequenced. By aligning with the miRNA database, four conserved miRNAs were finally obtained. Further analysis of their transcriptional expression revealed that all miRNAs were expressed at different stages of cotton ovule development. Among them, miR167a, miR172c and miR394b expressed differently in roots and leaves of cotton seedlings. The target genes of 55 miRNAs were predicted, and most of the target genes encoded transcription factors, metabolic enzymes and developmental related proteins, which further proved that miRNAs are involved in the regulation of the morphogenesis and development of cotton fiber.