目的 研究银柴胡Stellaria dichotoma不同组织部位甾醇类成分含量及基因转录水平差异，挖掘银柴胡甾醇类成分生物合成途径关键酶基因。方法 采用高效液相色谱法测定银柴胡根、茎、叶和花中甾醇类成分的含量，结合全长转录组测序与比较转录组学分析，通过加权基因共表达网络分析（weighted gene coexpression network analysis，WGCNA）挖掘甾醇类成分生物合成的关键酶基因，并利用RT-qPCR验证其表达水平。结果 银柴胡中甾醇类成分在根、茎、叶和花中含量差异显著。转录组测序组装获得372 483条高质量全长转录本序列，其中有128 646条全长转录本得到注释。差异基因分析结果显示，发现根、茎、叶、花4个组织共有的基因有43 354条，占基因总数的50.02%。根与花相比，差异表达基因总数为9 647条，根和叶相比，差异表达基因为12 143条，叶与茎相比，差异表达基因仅有388条。并从中鉴定出119个甾醇类成分合成的关键酶基因。进一步通过WGCNA鉴定到43个基因共表达网络模块，在MEblue模块中鉴定了NP_NY_transcript_168676 (FPPS)、NP_NY_transcript_335811(SQS)、NP_NY_transcript_44001(SQS)、NP_NY_transcript_246835 (CAS)、NP_NY_transcript_328932(CAS)和 NP_NY_transcript_ 183565(GPPS) 6个关键酶基因。对上述关键酶基因表达情况进行RT-qPCR验证，结果与转录组数据表达趋势一致。结论 揭示了银柴胡甾醇类成分合成的关键酶基因，为解析调控其有效成分生物合成途径及分子机制提供研究基础。

Objective To investigate the differences in sterol contents and gene transcription levels in different tissues of Stellaria dichotoma var. lanceolata, and explore key enzyme genes related to the biosynthetic pathway of sterols in this plant. Methods The contents of sterols in the roots, stems, leaves and flowers of S. dichotoma var. lanceolata was determined by high-performance liquid chromatography (HPLC). Combined with full-length transcriptome sequencing and comparative transcriptomic analysis, weighted gene coexpression network analysis (WGCNA) was used to mine key enzyme genes associated with sterol biosynthesis. The expression levels of these genes were further validated using RT-qPCR. Results The sterols contents in the roots, stems, leaves, and flowers of S. dichotoma var. lanceolata exhibited significant differences. A total of 372 483 high-quality full-length transcripts were obtained by transcriptome sequencing, of which 128 646 full-length transcripts were successfully annotated. Differential gene analysis revealed that 43 354 genes were expressed across the four tissues (roots, stems, leaves and flowers), accounting for 50.02% of all total genes. There were 9 647 differentially expressed genes (DEGs) between roots and flowers, 12 143 DEGs between roots and leaves, and only 388 DEGs between leaves and stems. A total of 119 key enzyme genes involved in sterol biosynthesis were identified, and 43 gene co-expression modules were identified by WGCNA. In the MEblue module, six key enzyme genes involved in sterol biosynthesis were identified, including NP_NY_transcript_168676 (FPPS), NP_NY_transcript_335811(SQS), NP_NY_transcript_44001(SQS), NP_NY_ transcript_246835 (CAS), NP_NY_transcript_328932(CAS) and NP_NY_transcript_ 183565(GPPS). The expression levels of the above-mentioned key enzyme genes were validated by RT-qPCR, and the results showed expression trends consistent with those obtained from the transcriptome data. Conclusion This study revealed the key enzyme genes involved in sterol biosynthesis in S. dichotoma L.var. lanceolate, providing a research basis for analyzing the biosynthetic pathways and molecular mechanisms that regulate its active ingredients.

R282.12

宁夏回族自治区高等学校科学研究项目（NYG2024299）