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作物学报

作物学报 ›› 2026, Vol. 52 ›› Issue (10): 2912-2926.doi: 10.3724/SP.J.1006.2026.65001

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蓝型油菜2种愈伤组织分化能力分子机制的转录组比较与解析

杨琴莉1(), 张晓玲1, 张丽贤1,2, 李红利1,2, 张换样1, 李换丽1, 孙彩红1,2, 李静1, 朱永红1, 孙璇1, 姚琳1, 王丹1, 上官小霞1,*()   

  1. 1 山西农业大学棉花研究所, 山西运城 044000
    2 山西农业大学农学院, 山西晋中 030800
  • 收稿日期:2026-01-01 接受日期:2026-07-15 出版日期:2026-10-12 网络出版日期:2026-07-21
  • 通讯作者: 上官小霞, E-mail: sgxx74@126.com
  • 作者简介:杨琴莉, E-mail: 739768392@qq.com
  • 基金资助:
    山西省科技重大专项计划揭榜挂帅项目(202201140601025-5-04);山西省运城市市级科技计划项目(YCKJ-2025046)

Comparative transcriptome analysis reveals the molecular mechanisms underlying differences in differentiation capacity between two distinct callus types in Brassica napus

Yang Qin-Li1(), Zhang Xiao-Ling1, Zhang Li-Xian1,2, Li Hong-Li1,2, Zhang Huan-Yang1, Li Huan-Li1, Sun Cai-Hong1,2, Li Jing1, Zhu Yong-Hong1, Sun Xuan1, Yao Lin1, Wang Dan1, Shang-Guan Xiao-Xia1,*()   

  1. 1 Institute of Cotton Research, Shanxi Agricultural University, Yuncheng 044000, Shanxi, China
    2 College of Agriculture, Shanxi Agricultural University, Jinzhong 030800, Shanxi, China
  • Received:2026-01-01 Accepted:2026-07-15 Published:2026-10-12 Published online:2026-07-21
  • Contact: Shang-Guan Xiao-Xia, E-mail: sgxx74@126.com
  • Supported by:
    ‘Unveiling the List and Taking Command’ Project of Shanxi Province Science and Technology Major Special Project(202201140601025-5-04);Yuncheng City Science and Technology Plan Project(YCKJ-2025046)

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摘要：

甘蓝型油菜遗传转化效率受其愈伤组织分化能力的严重制约, 且不同基因型间差异显著。本研究通过2个关键时间点的转录组分析, 解析导致分化能力差异的分子机制, 为遗传改良提供理论依据与基因资源。在20份不同基因型甘蓝型油菜中筛选出愈伤组织分化能力差异极显著的典型材料ZP10 (强)和ZP15 (弱), 对二者分化0 DAI (即诱导后0 d)和30 DAI的愈伤组织进行转录组测序与比较分析。差异表达基因(DEGs)筛选共鉴定出1915个基因。GO与KEGG富集分析显示, 这些基因显著富集于植物激素信号转导、光合作用-天线蛋白、光合器官组装及细胞壁生物合成等通路。结合加权基因共表达网络分析(WGCNA), 最终筛选出25个关键候选基因, 其表达模式与高分化能力密切相关。qRT-PCR结果证实了转录组数据的可靠性。综上, 甘蓝型油菜的高愈伤组织分化能力与一个由多基因协同调控的复杂网络有关, 涉及激素信号响应、光合系统初步建立及细胞壁重构等过程。本研究鉴定得到的关键基因为深入解析油菜再生分子机理及培育高转化效率基因型提供了重要靶点。

关键词: 甘蓝型油菜, 遗传转化, 愈伤组织, 分化能力, 转录组测序

Abstract:

The genetic transformation efficiency of Brassica napus is severely constrained by callus differentiation capacity, which varies greatly among genotypes. In this study, transcriptome analysis was performed at two key time points to elucidate the molecular mechanisms underlying differences in callus differentiation capacity, thereby providing a theoretical basis and genetic resources for genetic improvement. Two representative materials with highly significant differences in callus differentiation capacity, ZP10 with high differentiation capacity and ZP15 with low differentiation capacity, were selected from 20 B. napus genotypes. Transcriptome sequencing and comparative analysis were conducted on calli from these two lines at 0 and 30 days after induction (DAI). A total of 1915 genes were identified through differentially expressed gene (DEG) screening. GO and KEGG enrichment analyses showed that these genes were significantly enriched in pathways related to plant hormone signal transduction, photosynthesis-antenna proteins, photosynthetic organ assembly, and cell wall biosynthesis. Combined with weighted gene co-expression network analysis (WGCNA), 25 key candidate genes were ultimately identified, and their expression patterns were closely associated with high differentiation capacity. The reliability of the transcriptome data was confirmed by qRT-PCR. In conclusion, the high callus differentiation capacity of B. napus is associated with a complex regulatory network coordinated by multiple genes and involving hormone signaling responses, initial establishment of the photosynthetic system, and cell wall remodeling. The key genes identified in this study provide important targets for further elucidating the molecular mechanisms underlying rapeseed regeneration and for breeding genotypes with high transformation efficiency.

Key words: Brassica napus L., genetic transformation, callus, differentiation ability, RNA-Seq

表1

引物序列"

引物名称
Primer name		 引物序列
Primer sequence (5′-3′)
BnActin-F TGTGCCAATCTACGAGGGTTTC
BnActin-R TCTCACAATTTCCCGCTCGG
BnaC07G0469000ZS-F (ABI1-F) GACAACAACGGCGAGACTTC
BnaC07G0469000ZS-R (ABI1-R) TCCTCTCTCTACAATACTCCGCA
BnaA07G0340700ZS-F (EIL3-F) TCAGGCAACGAAGGAGGGGC
BnaA07G0340700ZS-R (EIL3-R) AGGAGGACTCTGGTTCTGCGGC
BnaA08G0267000ZS-F (CAP10B-F) GTGCTCAGTGGTCTTGGTCC
BnaA08G0267000ZS-R (CAP10B-R) GAGAAGGGACCCGAACGAG
BnaC02G0368500ZS-F (LHCB5-F) CACGCCAGATGGGCTATG
BnaC02G0368500ZS-R (LHCB5-R) AGAACAACCTCAGCAACTACAGC
BnaC04G0266800ZS-F (LHCA2-F) CCTGGTGACTTCGGGTTTGAT
BnaC04G0266800ZS-R (LHCA2-R) TTGTTGTTTGGGAAGATTGGGTC
BnaC04G0553100ZS-F (PAL1-F) TGTGAAAGCGAGTAGTGATTGGG
BnaC04G0553100ZS-R (PAL1-R) CTCTTGTGGCGGAGTGTGGTA

表2

不同油菜品系愈伤组织的诱导及分化情况"

材料编号
Material number		 接种外植体数
Number of inoculated explants		 愈伤组织的诱导率
Callus induction rate (%)		 出愈伤后接种外植体数
Number of explants
inoculated post-callus		 分化率
Callus differentiation
rate (%)		 再生苗率
Plant regeneration rate (%)
ZP1 60 47.78 def 30 47.22 bcdef 23.33 def
ZP2 60 43.33 defg 30 43.89 bcdefg 22.22 ef
ZP3 60 41.67 efg 30 40.00 defgh 20.00 fg
ZP4 60 55.00 cde 30 56.67 b 28.89 cde
ZP5 60 36.11 fg 30 36.11 efgh 18.89 fg
ZP6 60 41.67 efg 30 42.78 bcdefg 22.22 ef
ZP7 60 70.56 ab 30 72.22 a 37.78 b
ZP8 60 31.11 gh 30 32.22 gh 18.89 fg
ZP9 60 52.78 cde 30 55.56 bc 31.11 bcd
ZP10 60 80.56 a 30 80.00 a 46.67 a
ZP11 60 45.56 def 30 46.11 bcdefg 25.56 cdef
ZP12 60 53.89 cde 30 48.33 bcde 25.56 cdef
ZP13 60 21.67 h 30 38.89 efgh 13.33 g
ZP14 60 41.67 efg 30 41.67 cdefgh 24.44 cdef
ZP15 60 63.89 bc 30 27.78 h 20.00 fg
ZP16 60 47.22 def 30 45.00 bcdefg 22.22 ef
ZP17 60 34.44 fg 30 33.89 fgh 32.22 bc
ZP18 60 70.00 ab 30 69.44 a 36.67 b
ZP19 60 56.11 cd 30 53.89 bcd 26.67 cdef
ZP20 60 55.00 cde 30 56.11 b 26.67 cdef

图1

ZP10和ZP15不同分化时期愈伤组织形态 标尺: 培养皿中愈伤组织整体形态的标尺为1 cm; 愈伤组织显微形态的标尺为1 mm。DAI: 诱导后天数。"

表3

样本RNA测序数据产出及质控指标"

样本
Sample		 原始读数
Raw reads		 清洁读数
Clean reads		 清洁读段占比
Proportion of clean reads (%)		 Q20
(%)		 Q30
(%)		 平均GC含量
Mean GC content
(%)
YS10-1 44,413,342 43,995,642 99.06 98.87 96.46 46.72
YS10-2 39,924,584 39,504,914 98.95 98.81 96.31 46.67
YS10-3 43,630,348 43,304,644 99.25 98.84 96.42 46.80
YS15-1 48,312,106 48,011,976 99.38 98.97 96.78 46.88
YS15-2 43,486,404 43,233,720 99.42 98.85 96.51 46.73
YS15-3 43,992,790 43,732,868 99.41 98.89 96.61 46.81
FH10-1 41,487,832 41,156,180 99.20 98.77 96.25 46.78
FH10-2 43,749,892 43,398,526 99.20 98.83 96.42 46.87
FH10-3 39,741,766 39,332,990 98.97 98.91 96.65 46.94
FH15-1 41,207,924 40,916,276 99.29 98.86 96.48 46.98
FH15-2 40,900,106 40,570,336 99.19 98.76 96.13 46.98
FH15-3 39,623,008 39,298,456 99.18 98.87 96.47 47.04

图2

转录组数据的样本相关关系可视化 A: 主成分分析结果图; B: 样本间相关性热图。缩写同表3。"

图3

ZP10和ZP15不同阶段差异表达基因分析 A: 差异基因分组火山图; B: 差异表达基因聚类热图。缩写同表3。"

图4

差异基因重叠与表达模式的整合分析 A: 差异基因韦恩图; B: 差异表达基因聚类分析。缩写同表3。"

表4

ZP10愈伤组织中显著上调的转录因子家族及代表成员"

转录因子
家族
TF family		 上调的转录
因子数量
Number of up-regulated TFs		 代表性成员
Representative
member		 拟南芥同源
基因
Arabidopsis ortholog		 上调倍数
Fold change		 功能预测
Putative function
AP2/ERF 14 BnaC09G0490600ZS BBM1 9.40 参与锌离子或其他金属/小分子的跨膜运输过程
Involved in the transmembrane transport of zinc ions, other metal ions, or small molecules
ARR-B 2 BnaC09G0597600ZS gluA 10.39 一种定位于溶酶体的糖基水解酶
A lysosome‑localized glycosyl hydrolase
NAC 5 BnaC02G0167800ZS At1g02270 7.48 参与钙信号调控的核酸代谢或磷酸化过程
Involved in nucleic acid metabolism or phosphorylation processes regulated by calcium signaling
LBD 4 BnaC09G0523600ZS ZIFL1 8.65 在生长素运输和干旱胁迫响应中发挥关键作用
Play key roles in auxin transport and drought stress response
WRKY 7 BnaC02G0135100ZS EXO70B1 9.80 在囊泡运输和免疫信号传导中起作用
Play roles in vesicle transport and immune signaling
bHLH 13 BnaC09G0549300ZS MBR1 8.01 与MBR2协同作用, 调控茉莉酸信号通路
Acts synergistically with MBR2 to regulate the jasmonic acid signaling pathway
MYB 3 BnaA02G0407800ZS EXL2 12.02 在连接营养信号与植物生长调控中发挥作用
Involved in linking nutrient signals to plant growth regulation
bZIP 5 BnaC02G0155400ZS HSP90-2 8.07 参与植物生长发育、环境信号整合及免疫应答等
Involved in plant growth and development, environmental signal integration, and immune responses

图5

愈伤组织分化时期差异表达基因的GO富集分析"

图6

愈伤组织分化阶段差异表达基因KEGG富集分析 富集程度随富集得分的增大而增强; 点的大小反映了富集基因数量的多少; 点的红色越深, 富集显著性越高。"

图7

样品和模块间的相关性热图 缩写同表3。每个模块内上方数字表示相关系数r, 下方括号内数字表示P值。"

图8

候选基因在不同材料中的表达量热图 缩写同表3。颜色由蓝到红表示基因的相对表达水平由低到高。"

图9

差异表达基因的qRT-PCR验证 缩写同表3。数据表示平均值±标准差(n = 3)。柱子上方不同小写字母表示在P < 0.05水平差异显著。"

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