五指毛桃转录组及黄酮类化合物生物合成关键基因分析

Transcriptome Analysis of Ficus hirta Vahl and Expression Profiling of Key Genes Involved in Flavonoid Biosynthesis

五指毛桃含有香豆素类、黄酮类等化学成分,具有抗炎、抑菌、抗病毒和肿瘤的药用价值。但是,其转录组和功能基因组等分子水平的研究较少报道。本研究首次采用RNA-seq高通量测序技术对五指毛桃的根、茎和叶进行转录组测序,共获得104335条unigenes,平均长度为578 bp,有62142条unigenes被注释到Nr、GO、Swiss-Prot、KEGG和KOG等数据库中,根据同源序列比对发现五指毛桃与川桑的同源性最高。通过比较五指毛桃不同组织部位的转录组测序结果,发现Leaf_vs_Root、Root_vs_Stem、Stem_vs_Leaf分别有1363、624和1006个差异表达的基因在KEGG途径显著富集,主要富集在植物信号转导、植物MAPK信号通路、黄酮类生物合成等代谢途径。进一步分析参与植物黄酮类生物合成途径中的差异表达基因,发现有16个差异表达的基因参与该代谢途径,主要在叶和茎中高表达,运用RT-qPCR对黄酮类化合物生物合成途径中9个关键基因的表达量进行验证,发现定量结果与RNA-seq数据一致,大部分基因都在五指毛桃茎和叶片中高表达,因此五指毛桃幼苗黄酮类化合物主要积累在叶片和茎中,这与民间主要以根茎作为药用部位存在差异,主要原因可能是与五指毛桃生长年龄有关。以上的研究结果为五指毛桃分子生物学的研究奠定基础,并为其活性成分的生物合成调控、栽培技术等提供参考依据。

Ficus hirta Vahl contains coumarins, flavonoids and other chemical components, which have anti-inflammatory, bacteriostatic, antiviral and tumor medicinal value. However, the molecular research such as transcriptome and functional genome have less studied. Transcriptome sequencing was performed for the first time on different tissues of F. hirta Vahl using the RNA-seq high-throughput sequencing technique, and the result showed that a total of 10 4335 unigenes were obtained and 62 142 unigenes were annotated to the database of Nr, GO, Swiss-Prot, KEGG and KOG et al. F. hirta Vahl had the highest homology with Morus notabilis, according to the homologous sequence alignment. Comparative transcriptome analysis of different tissues of F. hirta Vahl showed that a total of 1363, 624 and 1006 genes were differentially expressed in the pairwise comparisons of LeafvsRoot, RootvsStem and StemvsLeaf, respectively. Among them, plant signal transduction, MAPK signal pathway-plant, and flavonoid biosynthesis related genes were significantly enriched, especially flavonoid biosynthesis pathway. Further analysis of differentially expressed genes involved in flavonoid biosynthesis showed that there were 16 differentially expressed genes involved in the pathway, which were mainly highly expressed in leaves and stems. The expression levels of key genes involved in flavonoid biosynthesis were verified by RT-qPCR, and the results were consistent with those of transcriptome sequencing. Therefore, flavonoids were mainly accumulated in the leaves and stems from young F. hirta Vahl. The above research results could lay a foundation for the research of molecular biology of F. hirta Vahl, and provide reference for the active component biosynthesis regulation and cultivation techniques.