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作物学报 ›› 2025, Vol. 51 ›› Issue (1): 134-148.doi: 10.3724/SP.J.1006.2025.44035

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蓝型油菜漆酶基因家族成员表达模式及与茎秆抗折力的关联分析

徐林珊1(), 郜耿东1, 王宇1, 王家星1, 杨吉招1, 武亚瑞1, 张宵寒1, 常影1, 李真1, 谢雄泽2, 龚德平3, 王晶1,*(), 葛贤宏1   

  1. 1华中农业大学植物科学技术学院, 湖北武汉 430070
    2襄阳市农业科学院, 湖北襄阳 441057
    3荆州农业科学院, 湖北荆州 434000
  • 收稿日期:2024-02-27 接受日期:2024-08-15 出版日期:2025-01-12 网络出版日期:2024-09-02
  • 通讯作者: *王晶, E-mail: wangjing@mail.hzau.edu.cn
  • 作者简介:E-mail: linshan@webmail.hzau.edu.cn
  • 基金资助:
    国家重点研发计划项目课题“多抗油菜新种质创制与应用”(2023YFD1201402);湖北省重点研发计划“耐密植超高产油菜品种高通量智能化选育”(2023BBB030)

Analysis of expression patterns of laccase gene family members in Brassica napus and their association with stem fracture resistance

XU Lin-Shan1(), GAO Geng-Dong1, WANG Yu1, WANG Jia-Xing1, YANG Ji-Zhao1, WU Ya-Rui1, ZHANG Xiao-Han1, CHANG Ying1, LI Zhen1, XIE Xiong-Ze2, GONG De-Ping3, WANG Jing1,*(), GE Xian-Hong1   

  1. 1College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, Hubei, China
    2Xiangyang Academy of Agricultural Sciences, Xiangyang 441057, Hubei, China
    3Jingzhou Academy of Agricultural Sciences, Jingzhou 434000, Hubei, China
  • Received:2024-02-27 Accepted:2024-08-15 Published:2025-01-12 Published online:2024-09-02
  • Contact: *E-mail: wangjing@mail.hzau.edu.cn
  • Supported by:
    National Key Research and Development Program ‘Creation and Application of New Germplasm of Multi-resistance Rapeseed’(2023YFD1201402);Hubei Key Research and Development Program ‘High-throughput Intelligent Selection and Breeding of High-yield Rapeseed Varieties with High Resistance to Dense Planting’(2023BBB030)

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摘要：

漆酶是一种含铜的多酚氧化酶家族, 在植物中主要参与木质素合成以及抵御各种逆境胁迫。本研究对甘蓝型油菜漆酶基因(BnaLACs)家族成员进行鉴定, 通过氨基酸数量、分子量、等电点、不稳定系数以及脂溶性系数等指标衡量其理化性质。后对其染色体位置、进化关系、基因结构、组织部位表达模式等进行预测和分析。结果表明, 甘蓝型油菜基因组共有53个BnaLACs家族成员, 基本为碱性、稳定蛋白, 大多数BnaLACs定位在液泡膜和细胞外。基因结构分析发现, BnaLACs结构较为保守。组织部位表达模式分析表明, 除花药外, BnaLACs在各组织部位均有表达, 其中在根、种子、角果皮和茎秆中表达量较高。分析茎秆中BnaLAC4s表达模式发现, BnaA05G0074200ZS与甘蓝型油菜抗倒性显著相关; 单倍型分析表明, 包含BnaA05G0074200ZS两种单倍型的品系间抗倒性、木质素含量均存在显著差异。研究结果将为进一步解析甘蓝型油菜漆酶基因家族功能及茎秆抗倒伏机制奠定基础。

关键词: 甘蓝型油菜, 漆酶, 进化, 表达模式, 单倍型, 抗倒性

Abstract:

Laccase is a family of copper-containing polyphenol oxidases primarily involved in lignin synthesis and resistance to various stresses in plants. In this study, members of the laccase gene family (BnaLACs) in Brassica napus were identified, and their physical and chemical properties were measured, including the number of amino acids, molecular weight, isoelectric point, instability index, and aliphatic index. The chromosome positions, evolutionary relationships, gene structures, and tissue expression patterns of these genes were subsequently predicted and analyzed. The results showed that the Brassica napus genome contains 53 BnaLAC family members, which are generally alkaline and stable proteins. Most BnaLACs are located in the vacuole membrane and outside the cell. Gene structure analysis revealed that BnaLACs have conserved structures. Tissue expression pattern analysis indicated that BnaLACs are expressed in all tissues except anthers, with higher expression levels in roots, seeds, silique walls, and stems. The expression pattern of BnaLAC4s in stems was specifically analyzed, and it was found that BnaA05G0074200ZS is significantly correlated with lodging resistance in Brassica napus. Haplotype analysis showed significant differences in lodging resistance and lignin content between different BnaA05G0074200ZS haplotype lines. The results of this study provide a foundation for further analysis of the functions of the laccase gene family in Brassica napus and the mechanisms underlying stem lodging resistance.

Key words: Brassica napus, laccase, evolution, expression pattern, haplotype, lodging resistance

表1

BnaA05G0074200ZS验证所用引物"

引物类型Primer type 引物名称Primer name 引物序列Primer sequence (5°-3°)
单倍型鉴定引物
Haplotype identification primer		 BnaLAC-A5-3329-F1 CTACTGTTGTTCGTCGTTTCTATTG
BnaLAC-A5-3329-R2 CTACTCACACACATTATTGTGGCT
BnaA5LAC-3K-F2 CTCCTCGGTTCTCTTTCTTCTA
BnaA5LAC-3K-R2 CATCTAATTGTATGTCTTATCGCCTTT
qRT-PCR引物
qRT-PCR primer		 qRTBnaLAC-A5F2 AGGCGCTTAAGTCTGGATTAGCCC
qRTBnaLAC-A5R2 AACGGTTTGACGTAGACAGCGTCC
BnENTH-F GTTTAGACCCGTTGCTGCTC
BnENTH-R TTGTCCATCTCAGCCATTTG

图1

BnaLACs在甘蓝型油菜染色体上的分布"

图2

拟南芥和甘蓝型油菜中LAC家族成员的系统发育树 共分为9类, 每一类用不同颜色标注。"

图3

拟南芥和甘蓝型油菜中LAC家族成员保守Motif以及基因结构分析 根据系统发育关系将BnaLAC家族共分为9类, 每一类用不同颜色标注。BnaLAC家族共鉴定到10种保守Motif, 不同颜色的方块表示不同Motif。基因结构中黄色方块表示外显子, 绿色方块表示UTR区域, 黑色线表示内含子。"

图4

甘蓝型油菜BnaLACs家族成员全生育期组织表达模式 各组织部位中BnaLACs表达量，组织部位分别为2 mm和4 mm花蕾，花丝，花瓣，花粉，花專，子叶，莲座叶，1~23片叶,根,开花后14~64d种子，开花后2~60d角果皮，下、中和上部"

图5

BnaLACs家族成员不同组织表达模式 A: 子叶、莲座叶以及1~23片叶中BnaLACs表达量。B: 花后14~64 d种子中BnaLACs表达量。C: 花后2~60 d角果皮中BnaLACs表达量。D: 下、中和上部茎秆中BnaLACs表达量。E: 根中BnaLACs表达量。F: 2 mm和4 mm花蕾中BnaLACs表达量。G: 花丝中BnaLACs表达量。"

图6

ZS11以及倒伏品种L23中 BnaLAC4s表达模式以及木质素含量 A、B、C为甘蓝型油菜数据库中ZS11表达模式。D、E、F为ZS11和L23中BnaLAC4s表达模式, ZS11: 中双11号。G: 木质素含量。t检验, *表示在0.05概率水平差异显著。"

图7

自然群体中BnaA05G0074200ZS单倍型分布情况 A: BnaA05G0074200ZS单倍型类型。B: 自然群体中BnaA05G0074200ZS单倍型与抗折力关系; t检验, P=0.016 < 0.05, 差异显著。C: 不同单倍型中BnaA05G0074200ZS相对表达量, C152和C344为hap1, C275和C474为hap2, 通过ANOVA方法分析表达差异显著性, 不同的小写字母代表在0.05概率水平差异显著。D: 不同单倍型木质素含量。"

表2

BnaLAC家族成员信息以及理化性质"

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