地黄microRNAs和靶基因的生物信息学预测及验证

Bioinformatic prediction and validation of conserved microRNAs and their target genes in Rehmannia glutinosa

目的拟利用地黄Rehmannia glutinosa EST数据通过生物信息学手段预测新的地黄microRNAs(miRNA),为今后地黄miRNA的生物学研究奠定基础。方法根据miRNA家族在不同物种中的保守性,将miRBase数据库中的已知植物miRNA与通过高通量测序获得的93 172条地黄EST序列进行同源比对,按照miRNA前体应具备的标准进行筛选。结果预测到分属于8个家族的8条潜在地黄miRNA序列,并通过实时荧光定量PCR对8条预测的miRNA进行了检测,证实8条miRNA在地黄中真实存在。随后利用软件对8条地黄miRNA的靶基因进行预测,发现其靶基因主要编码与地黄生长发育、代谢以及胁迫响应等过程相关的蛋白。结论新预测的地黄miRNA及其靶基因为今后研究它们在地黄中的生物学功能奠定了基础。

Objective In order to establish the material basis for future studies on the biological function of miRNAs, we aimed to predict novel miRNAs from EST sequences of Rehmannia glutinosa by using bioinformatic strategies. Methods Since most of the plant miRNAs were conserved in plant species, all plant miRNAs deposited in miRNase were aligned to the 93 172 EST sequences generated by next generation high-throughput RNA sequencing technology and the putative miRNA precursors were screened according to serious criteria. Results Eight novel rehmannia miRNAs were identified which belonged to eight different families that were further validated by real-time PCR analysis. Then the eight rehmannia miRNAs were subjected to target prediction analysis, and the results showed that the target genes encoded the proteins related to root growth, metabolism, stress responses and otherprocesses. Conclusion The miRNAs and their target genes identified in this study will provide clues to their biological functions in R. glutinosa.