基于转录组测序的襄麦冬块根甾体皂苷合成关联基因表达差异分析

Expression differences of steroidal saponin synthesis related genes in Liriope spicata var. prolifera Y. T. Ma via transciptome profiling

【目的】以襄麦冬(Liriope spicata var.prolifera Y. T. Ma)为对象,通过转录组学技术筛选、发掘相关功能基因,以期解析襄麦冬块根中甾体皂苷的生物合成。【方法】选取襄麦冬始见期块根样本和膨大期块根样本,使用转录组测序、序列注释分析、RT-qPCR等技术,验证并筛选多个与甾体皂苷合成相关的关键基因。【结果】共获得2个襄麦冬转录组的多条高质量序列(始见期样本59 405 731条;膨大期样本71 478 348条),共88 239条被注册到GO、COG、KOG、KEGG等数据库中。其中,通过GO数据库注释了42 388条拼接序列(分为3大类60个功能亚类),并在2个襄麦冬转录组中发现5563个差异表达基因;通过比对COG/KOG数据库,发现10 077条序列可注释到23个COG类群,8187条序列可注释到25个KOG类群,均与襄麦冬块根的生长及代谢有关。基于KEGG数据库的代谢通路分析表明,26 626条拼接序列可映射到不同的KEGG途径中,1981个差异表达基因被映射到227条KEGG途径;进一步筛选出43个可能参与甾体皂苷生物合成的差异表达基因后,并选取其中8个关键基因进行qRT-PCR验证。8个基因在襄麦冬不同生长时期表现出不同的表达量,与甾体皂苷合成有关的基因表达上调,且表达差异与转录组分析结果一致。该结果进一步验证了甾体皂苷在襄麦冬块茎中的生物合成途径,同时说明甾体皂苷的合成代谢与襄麦冬生长阶段性之间的相关性。【结论】初步揭示与襄麦冬甾体皂苷生物合成有潜在关联的基因和代谢通路,为今后进一步研究襄麦冬甾体皂苷的主要关联基因及代谢通路的功能以阐明其合成机理奠定了基础。

【Objective】The study aimed to analyze the biosynthesis of steroidal saponins in the root of Liriope spicata var. prolifera, and the related bio-synthesis genes were investigated.【Method】We used gene sequencing, transprome profiling and qRT-PCR technique to analyze the total RNA samples extracted from two different-staged L. spicata var. prolifera(first appearance period and expansion period), and several genes related to the bio-synthesis of steroidal saponins were indentified.【Result】Totally, we acquired high quality sequences after cDNA sequencing, there were 59 405 731 sequences acquired from the sample XMDSJQ(representing the root tuber sample in its first appearance period), and there were 71 478 348 sequences acquired from the sample XMDPDQ(representing the root tuber sample in its expansion period). After de novo assembly, the obtained sequences were respectively annotated databases, such as GO, COG, KOG, KEGG, et al. Compared with the GO database, a total number of 42 388 GO functional annotations were obtained, which were divided into 3 categories and 60 functional subcategories. There were 5563 differentially expressed genes. Among the annotated genes in the category of biological processes, the most genes were involved in cellular process. The KEGG database involved a total number of 227 pathways and cell cycle was the largest pathway group among the three main pathways. Also, 43 genes which might be involved in steroidal saponins biosynthesis were identified and 8 of them were quantified by qRT-PCR. The result showed that the biosynthsis of steroidal saponins from root tuber sample in the expansion period was more active, which was consistent with the profiling result of the transcriptome. 【Conclusion】The study preliminarily revealed the genes and metabolic pathways that were potentially related to the biosynthesis of steroidal saponins from L. spicata var. prolifera, and laid a foundation for further research on the functions of the main associated genes and metabolic pathways of steroidal saponins to clarify their synthesis mechanism.