摘要：

为解析杜鹃品种大鸳鸯锦花瓣粉色条纹形成的分子机制,挖掘与粉色条纹形成相关的关键基因,选取大鸳鸯锦盛花期花瓣,对花瓣白色区域与粉色条纹区域的色素分别进行测定,同时,利用 Illumina HiSeq^TM 测序平台对它们的转录组进行测序分析,并采用实时荧光定量 PCR(qRT-PCR)技术对测序结果进行验证。结果表明,大鸳鸯锦花瓣粉色条纹的形成是由花色苷积累引起的。花瓣粉色条纹区域与白色区域共鉴定出 7 086 个差异表达基因(DEGs),其中上调表达基因 3 802 个,下调表达基因 3 284 个。与花色苷形成相关的花青素生物合成、苯丙素生物合成和类黄酮生物合成途径在花瓣粉色条纹区域中更为活跃。根据 DEGs 的 GO 和 KEGG 富集分析结果,从与花色苷生物合成代谢相关及其调控途径共筛选出 31 个 DEGs,其中 26 个为花青素合成代谢结构基因, 5 个为转录调控基因(3 个 MYB 和 2 个 bHLH);26 个花青素合成代谢结构基因共编码 9 种酶,其中 20 个基因在花瓣粉色条纹组织中的表达水平明显高于白色花瓣组织;通过 NCBI 同源搜索发现 2 个 R2R3-MYB 和 1 个 bHLH 与已知调控果实或叶片中花青素合成有关的同源基因相似,它们在花瓣粉色条纹区域中的表达水平也均明显高于白色区域。选取10 个与花青素生物合成相关的差异表达基因,经 qRT-PCR 验证,发现基因表达趋势与转录组测序结果一致。

关键词: 杜鹃, 花瓣条纹, 花色苷, 转录组分析, 差异表达基因, 转录因子

Abstract:

In order to analyze the molecular mechanism of the formation of pink stripes on the petals of Dayuanyangjin,a Rhododendron cultivar,and to explore the key genes related to the formation of pink stripes,the petals of Dayuanyangjin in its full bloom period were selected to determine the pigments in the white and pink stripe regions of the petals,and at the same time,their transcriptome was sequenced and analyzed using Illumina HiSeq^TM sequencing platform.Sequencing results were verified by Real-time Fluorescent Quantitative PCR (qRT-PCR).The results showed that the formation of pink stripes on the petals of Dayuanyangjin was caused by the accumulation of anthocyanins.A total of 7 086 differentially expressed genes (DEGs) were identified in the pink striped and white regions of petals,of which 3 802 were up-regulated and 3 284 were down-regulated.The anthocyanin biosynthesis,phenylpropanoid biosynthesis and flavonoid biosynthesis pathways related to anthocyanin formation were more active in the pink stripe region of petals.According to the results of the GO and KEGG enrichment analysis of DEGs,31 DEGs were screened from the related and regulatory pathways of anthocyanin biosynthesis,of which 26 were structural genes of anthocyanin biosynthesis pathway and 5 were transcriptional regulation genes (3 MYB and 2 bHLH);26 anthocyanin anabolic structure genes encode 9 enzymes,and the expression levels of 20 genes in the pink stripe tissue of petals were significantly higher than those in the white petal tissue.Through NCBI homology search,it was found that two R2R3-MYB and one bHLH were similar to the known homologous genes related to the regulation of anthocyanin synthesis in fruits or leaves,and their expression levels in the pink stripe region of petals were also significantly higher than those in the white region.Ten differentially expressed genes related to anthocyanin biosynthesis were selected and verified by qRT-PCR.

Key words: Rhododendron, Petal stripe, Anthocyanins, Transcriptome analysis, Differential expression genes, Transcription factor