目的 从转录组层面对北柴胡Bupleurum chinense β-香树脂醇合酶（β-amyrin synthase，β-AS）基因家族成员进行鉴定，并对其表达及功能进行分析验证，以期为解析柴胡皂苷的合成和调控提供基础。方法 基于全长转录组数据挖掘北柴胡β-AS基因家族成员（BcBASs），利用在线软件对各基因进行生物信息学分析，利用大肠杆菌和烟草验证关键基因的功能。结果 从北柴胡转录组数据中挖掘到不冗余的β-AS基因6个，可分为3个亚家族，它们都具有典型保守区域DCTAE、MWCYCR和QW。进化分析表明，BcBAS1、BcBAS2、BcBAS4、BcBAS5、BcBAS6可能为单功能β-AS基因，BcBAS3可能为多功能β-AS基因，表达分析表明各基因在根和叶中的表达模式有差异。在大肠杆菌中成功表达了BcBAS1可溶性重组蛋白，相对分子质量约87 000，烟草瞬时表达表明BcBAS1编码蛋白具有β-AS功能，能促进β-香树脂醇的积累。结论 对北柴胡中6个不冗余的β-AS基因家族成员进行了鉴定和生物信息学分析，获得了BcBAS1可溶性重组蛋白，在烟草中验证了BcBAS1为有功能的β-AS，为柴胡皂苷合成调控机制和生物合成研究奠定了基础。

Objective To conduct identification analysis of the β-amyrin synthase (β-AS) gene family in Bupleurum chinense from the transcription level, and evaluate its expression and function for elucidating the biosynthetic pathways and regulatory mechanisms of saikosaponins (SSs). Methods Based on the transcriptomics data of B. chinense, the β-AS genes family (BcBASs) in B. chinense were mined, a series of tools were used for bioinformatics analysis and expression analysis of each gene, the key genes were transformed into Escherichia coli and tobacco for the functional validation. Results A total of six BcBASs members (BcBAS1— BcBAS6) were identified in this study, they were divided into three groups, all of them contained conserved regions including DCTAE, MWCYCR and QW. Evolutionary analyses indicated that BcBAS1, BcBAS2, BcBAS4, BcBAS5 and BcBAS6 may be monofunctional β-AS, BcBAS3 may be a multifunctional β-AS. Analysis of gene expression patterns showed that BcBASs were specifically expressed in different tissues. BcBAS1 was successfully expressed as solubl recombinant proteins in E. coli BL21(DE3), approximately 8.7 ×10⁴. The tobacco transient expression assay showed that BcBAS1 coded as β-AS, which affected the accumulation level of β-amyrin. Conclusion The members of β-AS gene family in B. chinense were identified, analyzed and validated. BcBAS1 soluble recombinant protein was obtained, and BcBAS1 was verified as a functional β-AS in tobacco. The results of this study will provide an important foundation for revealing the biosynthetic pathways and regulatory mechanisms of SSs.

R286.12

四川省“十四五”农作物育种攻关项目（2021YFYZ0012-13）；芸释医疗科技创新基金资助项目（20zh0234）