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作物学报

作物学报 ›› 2024, Vol. 50 ›› Issue (12): 3144-3154.doi: 10.3724/SP.J.1006.2024.43017

所属专题： 玉米：遗传育种·种质资源·分子遗传学

• 研究简报 • 上一篇    下一篇

玉米突变体caspl2b2的耐盐特性评价及转录组分析

张锦辉1(), 肖姿仪1, 李旭华3, 张明3, 贾春兰3, 潘振远2,*(), 邱法展1,*()   

  1. 1华中农业大学作物遗传改良全国重点实验室 / 湖北洪山实验室, 湖北武汉 430070
    2石河子大学农学院 / 绿洲生态农业兵团重点实验室, 新疆石河子 832003
    3山东登海种业股份有限公司, 山东莱州 261448
  • 收稿日期:2024-04-20 接受日期:2024-08-15 出版日期:2024-12-12 网络出版日期:2024-08-23
  • 通讯作者: *潘振远, E-mail: panzhenyuandawood@163.com; 邱法展, E-mail: qiufa
  • 作者简介:E-mail: 1907339567@qq.com
  • 基金资助:
    国家自然科学基金项目(U2106230);山东省重点研发计划项目(2022CXPT014)

Salt tolerance evaluation and transcriptome analysis of maize mutant caspl2b2

ZHANG Jin-Hui1(), XIAO Zi-Yi1, LI Xu-Hua3, ZHANG Ming3, JIA Chun-Lan3, PAN Zhen-Yuan2,*(), QIU Fa-Zhan1,*()   

  1. 1National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University / Hubei Hongshan Laboratory, Wuhan 430070, Hubei, China
    2College of Agriculture, Shihezi University / Key Laboratory of Oasis Ecological Agriculture Corps, Shihezi 832003, Xinjiang, China
    3Shandong Denghai Seed Industry Co., Ltd, Laizhou 261448, Shandong, China
  • Received:2024-04-20 Accepted:2024-08-15 Published:2024-12-12 Published online:2024-08-23
  • Contact: *E-mail: panzhenyuandawood@163.com; E-mail: qiufa
  • Supported by:
    National Natural Science Foundation of China(U2106230);Key Research and Development Program of Shandong Province(2022CXPT014)

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摘要：

盐胁迫敏感突变体是研究作物耐盐遗传基础和分子机制的重要遗传材料。本研究以自交系LY8405和突变体caspl2b2为材料, 开展正常生长、盐胁迫条件下苗期表型鉴定和生理生化指标测定分析。结果表明,与LY8405相比, 盐胁迫下caspl2b2存活率显著降低, 地上部分生长显著受到抑制; 地上部Na+离子含量显著升高, 丙二醛(MDA)含量显著升高; 蒸腾速率、气孔导度及胞间CO2浓度显著升高, 净光合速率显著下降。为揭示其盐胁迫下表型差异的分子基础, 对LY8405和caspl2b2突变体材料在正常生长和盐胁迫条件下叶片组织进行转录组分析。结果表明,差异表达基因主要富集到谷胱甘肽转移酶活性、谷胱甘肽代谢过程、氧化还原酶活性和细胞稳态等相关途径, 其中谷胱甘肽代谢过程最为显著。本研究不仅为作物耐盐遗传基础解析提供了重要的种质资源, 也为耐盐基因的挖掘及其遗传调控网络分析奠定了基础。

关键词: 玉米, 盐胁迫, 突变体, 转录组分析, 差异表达基因

Abstract:

Salt stress-sensitive mutants are crucial genetic materials for studying the genetic basis and molecular mechanisms of salt tolerance in crops. In this study, the maize inbred line LY8405 and the mutant caspl2b2 were used to investigate phenotype, physiological, and biochemical indices at the seedling stage under normal growth and salt stress conditions. The results showed that, compared to LY8405, the survival rate of caspl2b2 significantly decreased under salt stress, and the growth of aboveground parts was notably inhibited. The content of Na+ ions and malondialdehyde (MDA) in aboveground parts significantly increased, while transpiration rate, stomatal conductance, and intercellular CO2 concentration also increased significantly. Conversely, the net photosynthetic rate significantly decreased. To uncover the molecular basis of the phenotypic differences under salt stress, transcriptome analysis was performed on leaf tissues of LY8405 and caspl2b2 under normal growth and salt stress conditions. The results revealed that differentially expressed genes (DEGs) were mainly concentrated in glutathione transferase activity, glutathione metabolism, REDOX enzyme activity, and cell homeostasis, with glutathione metabolism being the most significant. This study not only provides important germplasm resources for the genetic basis analysis of crop salt tolerance, but also lays a foundation for the identification of salt tolerance genes and the analysis of genetic regulatory networks.

Key words: Zea mays L., salt stress, mutant, transcriptome analysis, differentially expressed genes

图1

caspl2b2突变体盐处理浓度筛选 A: 盐处理前WT和突变体caspl2b2的表型; B~D: 100 mmol L-1、150 mmol L-1和200 mmolL-1 NaCl处理7 d后WT和caspl2b2突变体的表型; E: 不同盐浓度处理下caspl2b2突变体的存活率。"

图2

盐处理后WT和caspl2b2突变体的存活率 A: 处理前WT和突变体caspl2b2的表型; B, D: 200 mmol L-1 NaCl处理3 d、7 d和14 d后的WT和caspl2b2突变体的表型; E: 盐处理不同天数后WT和caspl2b2突变体的存活率。"

图3

盐胁迫后 WT 和 caspl2b2 突变体的表型性状 A~D: 200 mmol L-1 NaCl处理后WT和caspl2b2突变体的苗高、初生根长、地上鲜重和地下鲜重的表型性状。E~G: 200 mmol L-1 NaCl处理3 d后WT和caspl2b2突变体的植株总体表型、叶片表型和根部表型。以上数据为平均值±标准差。Student’s t测验用于显著性分析。NS代表差异不显著。*代表P < 0.05, **代表P < 0.01。"

图4

盐胁迫后WT和caspl2b2突变体的Na+、K+含量 A: WT和caspl2b2突变体地上部Na+含量; B: K+含量; C: Na+ /K+; D: WT和caspl2b2突变体的MDA含量。以上数据为平均值±标准差。Student’s t测验用于显著性分析。NS代表差异不显著。*代表P < 0.05, **代表P < 0.01。"

图5

盐胁迫后WT和caspl2b2突变体的光合指标 A~D: 200 mmol L-1 NaCl处理3 d后WT和caspl2b2突变体的光合指标; A: 蒸腾速率; B: 气孔导度; C: 胞间CO2浓度; D: 净光合速率。以上数据为平均值±标准差。Student’s t测验用于显著性分析。NS代表差异不显著。*代表 P < 0.05, **代表 P < 0.01。"

表1

转录组测序相关数据"

样本
Sample		 测序总读数
Total reads number		 质控后读数
Clean reads number		 错误率
Error rate
(%)		 总比对率
Mapped ratio (%)		 Q20 碱基百分比
Q20 base percentage (%)		 Q30 碱基百分比
Q30 base percentage (%)
WT-1 21,635,978 20,957,262 0.02 96.86 98.22 94.71
WT-2 24,644,756 23,996,460 0.02 97.37 98.21 94.81
WT-3 22,386,955 21,537,487 0.02 96.21 98.12 94.55
Caspl2b2-1 25,381,514 24,489,459 0.02 96.49 98.20 94.72
Caspl2b2-2 23,662,504 22,826,123 0.02 96.47 98.14 94.68
Caspl2b2-3 23,551,867 22,839,156 0.02 96.97 97.94 94.32

图6

样本基因表达分布及相关性分析 A: 各样本基因表达分布; B: 样本间相关性。"

图7

差异基因火山图 Up: 上调基因; Down: 下调基因; Not significant: 无显著差异基因。"

图8

差异显著基因富集分析 A: 差异显著基因GO富集; B: 差异显著基因KEGG富集。"

表2

离子运输相关差异表达基因"

基因编号
Gene ID		 基因名称
Gene name		 描述
Description		 log2
(Fold Change)		 P值
P-value
Zm00001d003861 ZmHAK1 Potassium high-affinity transporter -0.15 0.44
Zm00001d042244 ZmHAK5 Potasium ion uptake permease 1 -0.61 0.55
Zm00001d049987 ZmHAK4 Probable potassium transporter 4 -0.31 0.71
Zm00001d040627 ZmHKT1 Sodium transporter HKT1 0.92 0.01
Zm00001d030955 ZmSOS3 CBL11; calcineurin B-like11: cbl11 -0.07 0.77
Zm00001d031232 ZmSOS1 Sodium/hydrogen exchanger 0.14 0.49
Zm00001d002428 ZmNSA1 EF hand family protein 0.46 3.00E-03
Zm00001d024587 ZmNPF6.4 Protein NRT1/PTR FAMILY 6.3 0.38 0.03

图9

qRT-PCR检测离子运输相关基因的表达 A: ZmHKT1基因的表达量; B: ZmHKT2基因的表达量; C: ZmHAK1基因的表达量; D: ZmHAK5基因的表达量。"

表3

凯氏带形成相关差异表达基因"

基因编号
Gene ID		 基因名称
Gene name		 描述
Description		 log2
(Fold Change)		 P值
P-value
Zm00001d038762 ZmRBOHF RBOH3; respiratory burst oxidase3 -0.32 0.11
Zm00001d027636 ZmESB1 Dirigent protein 16 0.42 0.87
Zm00001d040807 ZmCASPL1B2 CASPL4; CASP-like protein 4 0.10 0.60
Zm00001d005158 ZmCASPL1D1 CASPL3; CASP-like protein 3 -0.27 0.52
Zm00001d019457 ZmCASPL1D2 CASPL1D2; CASP-like protein 1D2 -0.44 0.05

表4

谷胱甘肽相关差异表达基因"

基因编号
Gene ID		 基因名称
Gene name		 描述
Description		 log2
(Fold Change)		 P值
P-value
Zm00001d049657 ZmGSTU1 GST26; glutathione transferase 26 -0.31 0.34
Zm00001d038192 ZmGSTU2 GST41; glutathione transferase 41 1.08 0.37
Zm00001d016860 ZmGSTU4 GST17; glutathione transferase 17 0.31 0.07
Zm00001d036951 ZmGSTU5 GST19; glutathione transferase 19 1.28 3.28E-09
Zm00001d043787 ZmGSTU6 GSTU6; glutathione S-transferase 6 0.85 0.003
Zm00001d043795 ZmGSTU10 GSTU10; glutathione S-transferase 10 0.98 8.47E-05
Zm00001d043789 ZmGSTU11 GSTU11; glutathione S-transferase 11 2.40 0.01
Zm00001d029706 ZmGSTU12 GST39; glutathione transferase 39 1.10 3.50E-07
Zm00001d043344 ZmGSTF3 GST8; glutathione transferase 8 0.80 0.20
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