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作物学报 ›› 2023, Vol. 49 ›› Issue (1): 62-72.doi: 10.3724/SP.J.1006.2023.24007

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

大豆生长节间响应温度和外源GA诱导的赤霉素途径关键基因分析

齐阳阳(), 窦汝娜, 赵彩桐, 张帜, 李文滨, 姜振峰()   

  1. 东北农业大学大豆生物学教育部重点实验室 / 农业农村部东北大豆生物学与遗传育种重点实验室, 黑龙江哈尔滨 150030
  • 收稿日期:2022-01-04 接受日期:2022-03-25 出版日期:2023-01-12 网络出版日期:2022-04-20
  • 通讯作者: 姜振峰
  • 作者简介:E-mail: qyy2022@126.com
  • 基金资助:
    国家自然科学基金项目(31571693);国家自然科学基金项目(32172072);财政部和农业农村部国家现代农业产业技术体系建设专项(大豆, 重点任务)(CARS-04-04B);黑龙江省自然科学基金项目(LH2021C025)

Analysis of key genes involved in GA pathway responding to temperature and exogenous GA related to internode development in soybean

QI Yang-Yang(), DOU Ru-Na, ZHAO Cai-Tong, ZHANG Zhi, LI Wen-Bin, JIANG Zhen-Feng()   

  1. Northeast Agricultural University, Key Laboratory of Soybean Biology in Chinese Ministry of Education / Key Laboratory of Soybean Biology and Genetics Breeding of Ministry of Agriculture and Rural Affairs, Harbin 150030, Heilongjiang, China
  • Received:2022-01-04 Accepted:2022-03-25 Published:2023-01-12 Published online:2022-04-20
  • Contact: JIANG Zhen-Feng
  • Supported by:
    National Natural Science Foundation of China(31571693);National Natural Science Foundation of China(32172072);China Agriculture Research System of MOF and MARA (Soybean, Key Task)(CARS-04-04B);Natural Science Foundation of Heilongjiang Province(LH2021C025)

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摘要：

为了解大豆节间在不同温度和外源赤霉素(gibberellic acid, GA)诱导条件下的表型变化规律, 分析GA合成重要途径和挖掘调控节间的关键候选基因, 本研究将大豆品种Charleston在培养箱条件和室外盆栽条件种植, 进行不同温度处理和外源GA涂抹处理, 利用徒手切片配合显微照相方法, 分析大豆节间长度和细胞形态变化; 利用液相色谱-质谱联机结合转录组测序方法分析大豆节间GA合成主要通路并挖掘调控节间生长的关键候选基因。结果表明在25℃和30℃条件下, 外源涂抹不同浓度GA可以诱导大豆生长节间伸长, 随着伸长量的增加, 大豆节间都变得纤细。外源GA对细胞作用效果主要为促进伸长, 对宽度影响不明显。高温处理对节间的伸长效果高于低温处理。本研究鉴定到GA2氧化酶基因在大豆生长节间表达和较高含量的GA19和GA53, 及这2种GA下游的GA20 (活性GA前体), 以及这条合成途径的活性GA产物GA3也被检测到都存在于细胞伸长区组织, 说明从GA前体物质到GA53, 再到GA19, 通过GA20最终合成GA3是大豆节间生长的一条重要GA合成通路, 进一步说明GA2氧化酶在大豆节间生长过程中有重要作用。挖掘到某些DELLA、GA和PIF基因家族成员具有组织表达特异性, 为调节大豆节间和株高提供了候选基因。

关键词: 大豆, 生长节间, GA途径, GA-GID1-DELLA

Abstract:

The objective of this study is to explore the effects of temperature and exogenous GA on the development of soybean internode and the key genes related to gibberellin (GA) signal transduction pathway. The soybean variety ‘Charleston’ was grown in chambers or in pots under outdoor condition, and subjected to the treatments with different temperature and concentration of GA solutions. Phenotype observation, section with hand, LC-MS analysis, and RNA-seq experiments were performed to investigate the internode changes and the genes involved in the GA signal transduction pathway. Different temperature and exogenous GA all induced the internode elongation and the longer the length increased, the slender the internode became. Exogenous GA had an evident elongating effect on the cell length of the internode despite of no effect on the cell width. The internodes grown in 30℃ growth condition were longer than those in 25℃ growth condition. GA2-oxidase, GA19, GA53, GA20, and bioactive GA3 were detected in the elongation zone of soybean internode, suggesting that GA2-oxidase might play an important role in the developmental process of soybean internode. Tissue-specific genes in DELLA, GAI, and PIF gene families could be identified and facilitate the gene selection to regulate the internode growth and plant height from GA signal transduction pathway. Soybean internode was regulated by temperature and exogenous GA. From GA53 to GA19 to GA20 and ultimately to GA3 was an important GA synthesis pathway. GA2-oxidase played an important role on the internode development of soybean. The GA-GID1-DELLA complex was expressed specifically in soybean internode. The candidate genes related to the internode development could be filtered from GID1 and DELLA gene family.

Key words: soybean, internode development, GA pathway, GA-GID1-DELLA

图1

温度升高促进对大豆节间生长 左侧2株为高温处理; 右侧2株低温处理。"

表1

不同温度对大豆生长节间的影响"

温度处理
Temperature treatment		 处理前
Before treatment (cm)		 12 h后
After 12 hours (cm)		 差值
Range (cm)
30℃ 3.02±0.32 3.38±0.35 0.36±0.03
25℃ 2.74±0.13 2.99±0.14 0.25±0.01

图2

外源GA促进大豆节间生长 A: 100 mmol L-1 GA处理; B: 250 mmol L-1 GA处理; C: 对照。"

图3

不同浓度GA外源施用对大豆节间生长影响 A: 25℃条件下GA处理; B: 30℃条件下GA处理。0 h: 未进行GA处理节间长度; 24 h: GA处理24 h后节间长度。"

图4

外源GA处理条件促进不同温度条件下大豆节间伸长区和成熟区细胞响应 A: 高温EZ; B: 高温MZ; C: 低温EZ; D: 低温MZ。"

表2

外源GA处理不同温度条件下大豆节间EZ和MZ细胞响应情况"

节间
Internode		 30℃ 25℃
长度 Length (pixel) 宽度 Width (pixel) 长度 Length (pixel) 宽度 Width (pixel)
伸长区 EZ 206.00±4.48 63.78±6.00 131.59±6.48 54.87±1.20
成熟区 MZ 708.33±21.20 65.78±1.11 665.79±27.51 79.18±2.51

表3

大豆植株生长节间不同GA含量"

节间
Internode		 GA3 GA15 GA19 GA20 GA53
伸长区 EZ 0.13±0.01 0.24±0.03 48.14±0.01 0.24±0.02 19.40±0.01
成熟区 MZ — — 47.91±0.02 — 19.38±0.02

图5

节间生长关键基因响应温度和外源GA的表达量变化 HCK: 30℃对照; LCK: 25℃对照; HG: 30℃ GA处理; LG: 25℃ GA处理。"

图6

高低密度条件下GA途径关键基因表达量变化 CD1: 低密度茎尖第1节; CD21: 低密度第2节伸长区; CD22: 低密度第2节成熟区; CD3: 低密度第3节; CH1: 高密度茎尖第1节; CH21: 高密度第2节伸长区; CH22: 高密度第2节成熟区; CH3: 高密度第3节。"

表4

GA信号转导关键基因的组织特异表达分析"

基因
Gene name		 嫩叶
Young leaf		 花
Flower		 1 cm荚
One-cm pod		 10 d种子
Seed of 10 DAF		 42 d种子
Seed of 42 DAF		 根
Root		 根瘤
Nodule
Glyma.12g213700
Glyma.13g288000 2.88 3.91 6.75
Glyma.20g141200 1.60 5.00 4.03
Glyma.05g130600 8.09
Glyma.13g361700 3.44 3.61 1.58 0.35 6.34 3.70
Glyma.15g012100 3.83 1.27 4.58 2.67 6.34 6.61
Glyma.04g150500 2.05 4.54 4.43 1.15 1.48 1.73 0.86
Glyma.06g213100 2.37 3.25 1.30 1.03 1.70 2.36 1.97
Glyma.06g213100 2.37 3.25 1.30 1.03 1.70 2.36 1.97
Glyma.11g216500 -2.88 -0.36 0.01 -0.03 -2.63 -2.33 -4.65
Glyma.18g040000 -2.31 -0.33 1.40 0.77 -3.97 -2.12 -4.03
Glyma.08g095800 -0.83 -1.01 -0.08 -0.87 -1.33 -1.96 -2.62
Glyma.20g200500 3.75 2.83 2.63 1.28 0.17 3.09 1.76
Glyma.05g140400 -1.51 -1.34 -0.84 -2.29 -1.65 -1.80 -1.94
Glyma.10g190200 3.66 3.14 2.00 1.03 -1.08 4.77 2.44
Glyma.15g141400 5.00 6.61
Glyma.03g148300 1.20 -1.75 0.65 -0.67 1.13 -1.49 -1.42
Glyma.10g022900 2.62 1.57 6.17 4.67 1.39 0.88 0.46
Glyma.02g151100 3.51 2.25 3.51 2.09 2.88 1.55 1.22
Glyma.20g230600 2.70 3.42 2.05 3.09 2.25 2.02 1.53
Glyma.10g158000 1.44 1.96 1.43 0.50 2.25 0.39 -0.10
Glyma.03g170300
Glyma.03g225000
Glyma.10g142600 3.66 2.54 3.85 -0.35 3.17 3.50 3.16
Glyma.20g091200 6.71 4.97 0.06
Glyma.02g282100 7.83 3.68 8.09
Glyma.U031209
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