Sunkar Ramanjulu, Zhou Xuefeng, Zheng Yun, Zhang Weixiong, Zhu Jian-Kang
Department of Biochemistry and Molecular Biology, Oklahoma State University, Stillwater, OK 74078, USA.
BMC Plant Biol. 2008 Feb 29;8:25. doi: 10.1186/1471-2229-8-25.
Small RNA-guided gene silencing at the transcriptional and post-transcriptional levels has emerged as an important mode of gene regulation in plants and animals. Thus far, conventional sequencing of small RNA libraries from rice led to the identification of most of the conserved miRNAs. Deep sequencing of small RNA libraries is an effective approach to uncover rare and lineage- and/or species-specific microRNAs (miRNAs) in any organism.
In order to identify new miRNAs and possibly abiotic-stress regulated small RNAs in rice, three small RNA libraries were constructed from control rice seedlings and seedlings exposed to drought or salt stress, and then subjected to pyrosequencing. A total of 58,781, 43,003 and 80,990 unique genome-matching small RNAs were obtained from the control, drought and salt stress libraries, respectively. Sequence analysis confirmed the expression of most of the conserved miRNAs in rice. Importantly, 23 new miRNAs mostly each derived from a unique locus in rice genome were identified. Six of the new miRNAs are conserved in other monocots. Additionally, we identified 40 candidate miRNAs. Allowing not more than 3 mis-matches between a miRNA and its target mRNA, we predicted 20 targets for 9 of the new miRNAs.
Deep sequencing proved to be an effective strategy that allowed the discovery of 23 low-abundance new miRNAs and 40 candidate miRNAs in rice.
转录水平和转录后水平上由小RNA引导的基因沉默已成为动植物基因调控的一种重要模式。到目前为止，对水稻小RNA文库进行常规测序已鉴定出了大多数保守的微小RNA（miRNA）。对小RNA文库进行深度测序是在任何生物体中发现稀有以及谱系和/或物种特异性微小RNA（miRNA）的有效方法。
为了鉴定水稻中新的miRNA以及可能受非生物胁迫调控的小RNA，我们从对照水稻幼苗以及遭受干旱或盐胁迫的幼苗中构建了三个小RNA文库，然后对其进行焦磷酸测序。分别从对照、干旱和盐胁迫文库中获得了58,781、43,003和80,990个独特的与基因组匹配的小RNA。序列分析证实了水稻中大多数保守miRNA的表达。重要的是，鉴定出了23个新的miRNA，它们大多各自源自水稻基因组中的一个独特位点。其中6个新的miRNA在其他单子叶植物中是保守的。此外，我们鉴定出了40个候选miRNA。在一个miRNA与其靶mRNA之间允许不超过3个错配的情况下，我们为9个新的miRNA预测了20个靶标。
深度测序被证明是一种有效的策略，可用于在水稻中发现23个低丰度新miRNA和40个候选miRNA。
