月季‘绿萼’花器官发育相关microRNA的鉴定及分析

利用高通量测序技术,构建了中国古老月季‘绿萼’(Rosa chinensis ‘Viridiflora’)和‘月月粉’(R.chinensis‘Old Blush’)花蕾期的microRNA(miRNA)文库,并对其进行了测序和序列分析。结果显示,在‘绿萼’文库中,鉴定到已知的miRNA成熟体39个,miRNA前体42个;预测到新的miRNA成熟体56个,前体57个。在‘月月粉’文库中,鉴定到已知RNA成熟体39个,已知miRNA前体40个;预测到新的miRNA成熟体53个,前体57个。与‘月月粉’相比,‘绿萼’中差异表达的miRNA有31个,其中17个上调、14个下调。荧光定量PCR实验结果表明,miR156、miR398和miR535在2种月季的花蕾期表达上调,而miR167、miR172和miR396表达下调。进一步检测miR172和miR156在2种月季不同花器官中的表达差异,发现miR172在‘绿萼’的花瓣、雌、雄蕊中表达显著下调,提示miR172可能通过负调控其靶基因RcAP2的表达,在‘绿萼’花器官发育过程中起重要作用。

Identification of microRNA associated with flower organ development in Rosa chinensis ‘Viridiflora’

In this study, two independent microRNA (miRNA) libraries of Rosa chinensis ‘Viridiflora’and ‘Old Blush’ at the flower bud development stage were constructed and sequenced with Illumina sequencing. In total, 39 known miRNAs and 42 known pre-miRNAs were discovered, and 56 novel miRNAs and 57 novel pre-miRNAs were predicted in the ‘Viridiflora’library. In total, 39 known miRNAs and 40 known pre-miRNAs were identified, and 53 novel miRNAs and 57 novel pre-miRNAs were predicted in the ‘Old Blush’ library. Compared with ‘Old Blush’, there were 31 differentially expressed miRNAs in ‘Viridiflora’, with 17 up-regulated and 14 down-regulated. The RT-qPCR results showed that the expressions of miR156, miR398, and miR535 were up-regulated, whereas those of miR167, miR172, and miR396 were down-regulated in ‘Viridiflora’, identical to those obtained by sequencing. The expressions of miR172 and miR156 in different floral organs of the two rose species were detected by RT-qPCR. We found that miR172 was down-regulated in the petals, pistils, and stamens of ‘Viridiflora’. Furthermore, RcAP2 was previously reported to be up-regulated in relative flower organs, suggesting that miR172 may negatively regulate the expression of its target gene RcAP2. Thus, miR172 may play an important role in the development of floral organs of ‘Viridiflora’.