[目的]克隆灰毡毛忍冬MADS-box家族基因SOC1(SUPPRESSOR OF OVEREXPRESSION OF CO 1)，对其进行生物信息学和表达模式分析，为探究灰毡毛忍冬花冠不开放机制奠定理论基础。[方法]基于花蕾型灰毡毛忍冬转录组数据，筛选到开花调控基因LmSOC1 的Unigene序列并克隆全长cDNA序列，通过生物信息学分析和实时荧光定量PCR技术分析编码蛋白特性和不同品种中LmSOC1 基因的表达特异性，最后将LmSOC1基因插入到原核表达质粒载体pTOPO-D1 中，并在表达宿主BL21(DE3)中进行蛋白表达。[结果]LmSOC1基因包含645 bp的ORF区，LmSOC1蛋白不含信号肽、无跨膜区，为稳定的亲水性蛋白。系统进化树分析表明，LmSOC1 蛋白与黄花蒿、榴莲、可可等的SOC1 蛋白亲缘关系更近。实时荧光定量PCR分析表明，在 2 个花蕾型灰毡毛忍冬品种的花蕾和叶片中均检测到LmSOC1 基因的表达，而叶片中表达量明显高于花蕾。同时，在早期花蕾中LmSOC1 基因的表达量高于晚期花蕾，在'金翠蕾'品种中这一表达差异极显著(P＜0。01)。最后成功在大肠杆菌BL21(DE3)中表达出重组蛋白。[结论]通过对LmSOC1 基因的克隆和表达分析发现，该基因可能在灰毡毛忍冬花器官发育过程中发挥重要作用，为进一步研究该基因在植物花发育中的生物学功能奠定了基础。
Cloning and expression analysis of LmSOC1 gene related to floral organ development in Lonicera macranthoides
[Objective]SUPPRESOR OF OVEREXPRESSION OF CO 1(SOC1)gene from MADS-box family of Lonicera macranthoides was cloned and its bioinformatics and expression patterns were analyzed to lay a theoretical foundation for exploring the mechanism of corolla non-opening in L.macranthoides.[Method]In order to explore the functions of SOC1 genes in the flowering of L.macranthoides,this study screened and cloned the cDNA sequences of LmSOC1 genes from the transcriptome data of bud-type L.macranthoides.Bioinformatics analysis was used to analyze the characteristics of the encoded proteins,and quantitative reverse-transcription polymerase chain reaction to detect the specific expression of LmSOC1 in different varieties.And then the LmSOC1 gene extracted from L.macranthoides was successfully integrated into the prokaryotic expression plasmid pTOPO-D1.Subsequently,this recombinant plasmid was introduced into Escherichia coli strain BL21(DE3)to facilitate the expression of the target protein.[Result]The LmSOC1 gene harbored a 645 bp open reading frame(ORF),yielding an encoded protein characterized by stability and hydrophilicity,devoid of both transmembrane regions and signal peptides.Phylogenetic tree analysis proved that LmSOC1 protein had close genetic relationship with the LmSOC1 proteins from Artemisia annua,Durio zibethinus,and Theobroma cacao.The real-time fluorescence quantitative PCR analysis revealed the presence of LmSOC1 gene expression in both the buds and leaves of two bud type varieties of Lonicera japonica.Interestingly,the expression level was notably higher in the leaves compared to the buds.Furthermore,within the buds,the expression of the LmSOC1 gene was found to be higher in early flower buds compared to late flower buds.This discrepancy in expression was particularly significant in the'Jincuilei'variety,with a p-value of less than 0.01,indicating an extremely significant difference.Finally,the recombinant protein was successfully expressed in E.coli BL21(DE3).[Conclusion]Through cloning and expression analysis of the LmSOC1 gene,it was found that this gene may play an important role in the development of organs in L.macranthoides,and it laid a foundation for further research on the biological function of this gene.
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