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作物学报 ›› 2025, Vol. 51 ›› Issue (10): 2693-2704.doi: 10.3724/SP.J.1006.2025.55008

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

基于转录组分析鉴定甘蓝型油菜开花候选基因以及BnaCOR27功能验证

王晨1,2(), 贺丹1,2, 姚敏1,2, 邱萍1,2, 何昕1,2, 熊兴华1,2, 康雷1,2, 刘忠松1,2, 钱论文1,2,*()   

  1. 1湖南农业大学农学院, 湖南长沙 410128
    2岳麓山实验室, 湖南长沙 410082
  • 收稿日期:2025-01-16 接受日期:2025-07-09 出版日期:2025-10-12 网络出版日期:2025-07-22
  • 通讯作者: *钱论文, E-mail: qianlunwen@163.com
  • 作者简介:E-mail: 1048162651@qq.com
  • 基金资助:
    湖南省科技创新领军人才项目(2023RC1063);长沙市杰出创新青年培养计划项目(kq2107016)

Transcriptome-based identification of flowering candidate genes and functional characterization of BnaCOR27 in Brassica napus

WANG Chen1,2(), HE Dan1,2, YAO Min1,2, QIU Ping1,2, HE Xin1,2, XIONG Xing-Hua1,2, KANG Lei1,2, LIU Zhong-Song1,2, QIAN Lun-Wen1,2,*()   

  1. 1College of Agronomy, Hunan Agricultural University, Changsha 410128, Hunan, China
    2Yuelu Mountain Laboratory, Changsha 410082, Hunan, China
  • Received:2025-01-16 Accepted:2025-07-09 Published:2025-10-12 Published online:2025-07-22
  • Contact: *E-mail: qianlunwen@163.com
  • Supported by:
    Hunan Province Science and Technology Innovation Leading Talent Project(2023RC1063);Training Program for Excellent Young Innovators of Changsha(kq2107016)

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摘要：

开花是高等植物从营养生长到生殖生长的一个重要阶段, 影响生物量和种子产量。为了揭示甘蓝型油菜春化的调控网络以及挖掘重要候选基因, 本研究对7个甘蓝型油菜品种春化前后的叶片转录组数据进行分析, 鉴定出1305个差异表达基因, 其中上调基因有554个, 下调基因有751个。GO富集分析发现, 差异基因主要富集在春化途径、光周期途径、昼夜节律、花发育以及冷响应等生物学过程中, 进一步分析发现96个开花基因存在差异表达, 包括BnaVIN3、BnaFLC、BnaCOR27等基因。此外, 全基因组关联分析鉴定BnaCOR27-C04与开花时间显著关联, 结合加权基因共表达网络分析, 发现BnaCOR27与BnaFLC、BnaFT、BnaVIN3等基因直接相连形成潜在的网络调控甘蓝型油菜开花时间。进一步利用CRISPR/CAS9基因编辑技术对BnaCOR27进行敲除, 通过对T3代转基因植株表型分析, 发现敲除突变体株系表现出早花表型。这一结果能对已有的甘蓝型油菜开花研究进行补充和丰富, 为进一步改良开花性状提供遗传基础。

关键词: 甘蓝型油菜, 开花, 转录组分析, GWAS, BnaCOR27

Abstract:

Flowering is a critical developmental transition in higher plants, marking the shift from vegetative to reproductive growth, and it directly influences biomass accumulation and seed yield. To elucidate the regulatory network underlying vernalization in Brassica napus and identify key candidate genes, we analyzed the transcriptomes of leaves from seven rapeseed varieties before and after vernalization. A total of 1305 differentially expressed genes (DEGs) were identified, including 554 upregulated and 751 downregulated genes. GO enrichment analysis showed that these DEGs were primarily involved in biological processes such as the vernalization pathway, photoperiod pathway, circadian rhythm, flower development, and response to cold. Further analysis revealed differential expression in 96 flowering-related genes, including BnaVIN3, BnaFLC, BnaCOR27, among others. Additionally, genome-wide association studies (GWAS) indicated that BnaCOR27-C04 is significantly associated with flowering time. Integration with weighted gene co-expression network analysis (WGCNA) suggested that BnaCOR27 may interact with genes such as BnaFLC, BnaFT, and BnaVIN3, forming a potential regulatory network controlling flowering time in rapeseed. Furthermore, CRISPR/Cas9-mediated knockout of BnaCOR27 resulted in an early-flowering phenotype in T3 transgenic plants. These findings enhance our understanding of flowering regulation in B. napus and provide a genetic foundation for improving flowering-related traits in future breeding efforts.

Key words: Brassica napus, flowering, transcriptome analysis, GWAS, BnaCOR27

图1

差异基因火山图 纵坐标代表差异表达显著性, 横坐标代表基因表达量变化倍数, 红色点代表上调差异基因, 蓝色点代表下调差异基因。"

图2

差异表达基因GO富集分析 A: 上调基因GO富集分析; B: 下调基因GO富集分析。"

图3

热图显示与开花相关的差异表达基因的表达量 A: 上调差异基因的表达量热图; B: 下调差异基因的表达量热图。"

图4

显著相关区域候选基因的分析 A: 全基因组关联分析检测到单体型区域(16,048,376~16,594,153 bp)与开花期显著关联。红点表示BnaCOR27-C04中的SNP与开花期关联, 虚线表示显著性阈值; B: BnaCOR27-C04基因区域2个等位基因对应的表型分析。通过t检验分析等位基因座之间的差异。"

图5

共表达网络分析 A: 模块系统树图; B: 模块与春化相关性; C: 基因网络图。根据功能标注, 网络分为春化途径(粉色节点)、光周期途径(橙色节点)、花发育相关(淡紫色节点)、激素途径(深绿色节点)、低温途径(蓝绿色节点)、昼夜节律途径(翠绿色节点)、自主途径(紫色节点)和其他与开花有关的基因(灰色节点)。"

图6

转基因植株靶点突变序列分析及开花表型 A: BnaCOR27在A09染色体的敲除载体以及靶点突变情况示意图; B: BnaCOR27在C09染色体的敲除载体以及靶点突变情况示意图; C: 敲除突变体植株CAS:BnaCOR27-T3及对照植株ZS6开花时间表型图片; D: 敲除突变株系CAS:BnaCOR27-T3及对照植株ZS6开花时间柱状图; **: P < 0.01。在A09染色体上出现大片段缺失, C09染色体上插入单碱基。敲除突变体植株CAS:BnaCOR27-T3相较于对照植株ZS6开花时间提前约6.54 d。ZS6: 中双6号。标尺为5 cm。"

图7

CAS:BnaCOR27-T3转基因植株转录组差异基因分析 A: CAS:BnaCOR27-T3转基因材料差异基因火山图; B: CAS:BnaCOR27-T3转基因材料GO富集分析; C: CAS:BnaCOR27-T3转基因材料转录组差异基因表达量分析。ZS6: 中双6号。"

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