木薯等大戟科植物耐寒抗旱相关miRNA的分离与鉴定
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摘要
microRNA(miRNA)是一类～21nt的,在转录后水平上调节mRNA的非编码RNA。miRNA的调控作用涉及几乎所有重要的生物学过程。为了解miRNA在重要显花植物家族大戟科中的作用,本研究运用计算预测和实验分析相结合的方法,克服了多数大戟科物种缺乏基因组信息的问题,开展了在基因组水平上的大规模发掘和分析大戟科miRNA功能的系统研究。本研究利用已经释放的蓖麻基因组序列信息在蓖麻中预测了23个miRNA家族的85个miRNA,随后利用蓖麻(Ricinus communis),木薯(Manihot esculenta),橡胶树(Hevea brasiliensis)和麻风树(Jatropha curcas)这4个物种的苗期样品对这85个miRNA进行差异表达分析,发现其中58个miRNA(占85 miRNA的68.2%)至少在4个大戟科物种之一中表达,进一步研究发现48个miRNA(85 miRNA的56.5%)在低温和干旱胁迫条件下,以及在胁迫后的回复过程中有明显表达。以上研究结果表明miRNA具有物种和环境特异性的表达模式。另外,本研究利用Hitsensor软件,预测出63个miRNA与83个靶基因之间的258对互作,并对其中10个互作对进行了验证,证实了其中的6个互作对,并确定了靶基因的剪切位点。本研究是第一次在大戟科植物中收集miRNA及其靶基因,这些研究成果为今后在大戟科植物家族中研究生物和非生物胁迫奠定了基础。
MicroRNA(miRNA) is a class of～21nt non-coding RNAs that regulate mRNAs at the post-transcriptional level,miRNAs are key regulators of nearly all essential biological processes.Aiming at understanding miRNAs' functions in Euphorbiaceae, a large flowering plant family,we performed a genome-scale systematic study of miRNAs in Euphorbiaceae,by combining computational prediction and experimental analysis to overcome the difficulty of lack of genomes for most Euphorbiaceous species.Specifically,we predicted 85 conserved miRNAs of 23 miRNA families in Castor bean,and then experimentally verified and characterized 58(68.2%) of the 85 miRNAs in at least one of four agri-economically important Euphorbiaceous species, Castor bean(Ricinus communis),Cassava(Manihot esculenta),Rubber tree(Hevea brasiliensis),and Jatropha(Jatropha cureas),during normal seedling development. To elucidate their functions in stress response,we further verified and profiled 48 (56.5%) of the 85 miRNAs under cold and drought stresses as well as during the processes of stress recovery.The results revealed speciesandcondition-specific miRNA expression patterns.Finally,we predicted 258 miRNA:target binding partners, and identified the cleavage sites of six out of ten miRNA targets by modified 5' RACE.This study produced the first collection of miRNAs and their targets in Euphorbiaceae.Our results revealed wide conservation of many miRNAs and their diverse functions in Euphorbiaceous plants during seedling growth and in response to abiotic stresses.
MicroRNA(miRNA) is a class of～21nt non-coding RNAs that regulate mRNAs at the post-transcriptional level,miRNAs are key regulators of nearly all essential biological processes.Aiming at understanding miRNAs' functions in Euphorbiaceae, a large flowering plant family,we performed a genome-scale systematic study of miRNAs in Euphorbiaceae,by combining computational prediction and experimental analysis to overcome the difficulty of lack of genomes for most Euphorbiaceous species.Specifically,we predicted 85 conserved miRNAs of 23 miRNA families in Castor bean,and then experimentally verified and characterized 58(68.2%) of the 85 miRNAs in at least one of four agri-economically important Euphorbiaceous species, Castor bean(Ricinus communis),Cassava(Manihot esculenta),Rubber tree(Hevea brasiliensis),and Jatropha(Jatropha cureas),during normal seedling development. To elucidate their functions in stress response,we further verified and profiled 48 (56.5%) of the 85 miRNAs under cold and drought stresses as well as during the processes of stress recovery.The results revealed speciesandcondition-specific miRNA expression patterns.Finally,we predicted 258 miRNA:target binding partners, and identified the cleavage sites of six out of ten miRNA targets by modified 5' RACE.This study produced the first collection of miRNAs and their targets in Euphorbiaceae.Our results revealed wide conservation of many miRNAs and their diverse functions in Euphorbiaceous plants during seedling growth and in response to abiotic stresses.
引文
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