Identification of drought-responsive microRNAs in Medicago truncatula by genome-wide high-throughput sequencing

Two hundreds eighty three and 293 known miRNAs were identified from the control and drought stress libraries, respectively. In addition, 253 potential candidate miRNAs were identified, and among them 48 novel miRNAs/ novel members of known miRNA families whose complementary miRNAs were also detected. Both high-throughput sequencing and RT-qPCR confirmed that 22 members of 4 miRNA families were up-regulated and 10 members of 6 miRNA families were down-regulated in response to drought stress. Among the 48 novel miRNAs, 24 miRNAs were responsive to drought stress with 21 and 3 miRNAs being up- and down-regulated, respectively. The known and predicted targets of these miRNAs in response to drought stress are involved in diverse cellular processes in plants, such as development, transcription, protein degradation, detoxification, nutrient status and cross adaptation. To identify miRNAs in response to drought stress in M. truncatula, control and drought stress shoots were used to construct small RNA libraries, respectively. The date from Solexa sequencer (Illumina) was compared to identify miRNAs in response to drought stress.

本研究分别从对照文库与干旱胁迫文库中鉴定出283个和293个已知microRNA（miRNA）。此外，还鉴定出253个潜在候选miRNA，其中48个为新型miRNA或已知miRNA家族的新型成员，且同步检测到了其互补miRNA。高通量测序与实时荧光定量PCR（RT-qPCR）均证实，4个miRNA家族的22个成员在干旱胁迫下呈上调表达，6个miRNA家族的10个成员则呈下调表达。在这48个新型miRNA中，有24个miRNA对干旱胁迫产生响应，其中21个上调、3个下调。这些响应干旱胁迫的miRNA的已知靶标与预测靶标，参与植物体内多种细胞生物学过程，包括生长发育、转录调控、蛋白质降解、解毒作用、营养稳态维持以及交叉适应等。为鉴定蒺藜苜蓿（Medicago truncatula）中响应干旱胁迫的miRNA，研究人员分别以对照与干旱胁迫处理的地上组织为材料构建了小RNA文库。通过比对Illumina Solexa测序仪产生的测序数据，最终筛选得到响应干旱胁迫的miRNA。