生物技术通报 ›› 2023, Vol. 39 ›› Issue (10): 148-162.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0183
白苗1(), 田雯青1, 武帅1, 王敏1, 王利祥1, 岳爱琴1, 牛景萍3, 张永坡2, 高春艳2, 张武霞2, 郭数进1, 杜维俊1(), 赵晋忠2()
收稿日期:
2023-03-03
出版日期:
2023-10-26
发布日期:
2023-11-28
通讯作者:
杜维俊,女,博士,研究方向:作物遗传育种;E-mail: duweijun68@126.com;作者简介:
白苗,女,硕士,研究方向:大豆遗传育种;E-mail: 629901582@qq.com
基金资助:
BAI Miao1(), TIAN Wen-qing1, WU Shuai1, WANG Min1, WANG Li-xiang1, YUE Ai-qin1, NIU Jing-ping3, ZHANG Yong-po2, GAO Chun-yan2, ZHANG Wu-xia2, GUO Shu-jin1, DU Wei-jun1(), ZHAO Jin-zhong2()
Received:
2023-03-03
Published:
2023-10-26
Online:
2023-11-28
摘要:
明确大豆维生素组分含量及合成相关基因γ-TMT对激素和逆境胁迫的响应,为维生素E参与逆境胁迫研究开辟新思路。以山西省主栽品种晋大88为试验材料,在激素、逆境胁迫条件下对不同组织的维生素E组分含量、生理指标、γ-TMT表达模式进行分析。结果表明,维生素E主要存在于叶中,茎中有少量,根中不存在。在激素及盐等胁迫条件下,叶和茎的总维生素E含量小于对照,但γ-生育酚含量在NaCl、H2O2、ABA胁迫下含量高于对照。在激素和逆境胁迫环境下,大豆植株的生长被显著抑制,株高、鲜重、干重、SPAD值以及含水量皆小于对照。基因组数据挖掘到3个γ-TMT,启动子中均存在许多胁迫应答和激素响应元件,在激素和逆境胁迫下,γ-TMT1-γ-TMT3在根、叶中的表达均发生上调或下调。在NaCl胁迫条件下,大豆叶中γ-生育酚含量增加,而γ-TMT2、γ-TMT3的表达明显下调,推测γ-TMT2和γ-TMT3可能影响盐胁迫条件下维生素组分含量。激素和逆境胁迫显著抑制大豆茎和叶中维生素E的含量,进而抑制植株的生长,影响了大豆γ-TMT表达和维生素E组分。
白苗, 田雯青, 武帅, 王敏, 王利祥, 岳爱琴, 牛景萍, 张永坡, 高春艳, 张武霞, 郭数进, 杜维俊, 赵晋忠. 激素和逆境胁迫对大豆维生素E和γ-TMT表达的影响[J]. 生物技术通报, 2023, 39(10): 148-162.
BAI Miao, TIAN Wen-qing, WU Shuai, WANG Min, WANG Li-xiang, YUE Ai-qin, NIU Jing-ping, ZHANG Yong-po, GAO Chun-yan, ZHANG Wu-xia, GUO Shu-jin, DU Wei-jun, ZHAO Jin-zhong. Effects of Hormonal and Adversely Stress on Vitamin E and γ-TMT Gene Expression in Soybeans[J]. Biotechnology Bulletin, 2023, 39(10): 148-162.
基因名称 Gene name | 引物 Primer | 序列 Sequence(5'-3') |
---|---|---|
γ-TMT1 | F | GTGTGGTCCATGGAGAGTGG |
R | CCCTGTGGCACCATGTTACT | |
γ-TMT2 | F | CTGGCCGTCAGAGAATGGTT |
R | GGAGCAACCAGTGTAGGCAT | |
γ-TMT3 | F | TGACGCCCTAAAGCAACCAT |
R | CCCTGTGGCACCATGTTACT | |
GmCYP2 | F | CGGGACCAGTGTGCTTCTTCA |
R | GTGCTGCTACCCGAGCTAAT |
表1 γ-TMT的RT-qPCR扩增引物
Table 1 Primers for RT-qPCR amplification of γ-TMT gene
基因名称 Gene name | 引物 Primer | 序列 Sequence(5'-3') |
---|---|---|
γ-TMT1 | F | GTGTGGTCCATGGAGAGTGG |
R | CCCTGTGGCACCATGTTACT | |
γ-TMT2 | F | CTGGCCGTCAGAGAATGGTT |
R | GGAGCAACCAGTGTAGGCAT | |
γ-TMT3 | F | TGACGCCCTAAAGCAACCAT |
R | CCCTGTGGCACCATGTTACT | |
GmCYP2 | F | CGGGACCAGTGTGCTTCTTCA |
R | GTGCTGCTACCCGAGCTAAT |
图1 γ-TMT氨基酸序列的比对 A:丙氨酸;R:精氨酸;N:天冬酰胺;D:天冬氨酸;C:半胱氨酸;Q:谷氨酰胺;E:谷氨酸;G.甘氨酸;H:组氨酸;I:异亮氨酸;L:亮氨酸;K:赖氨酸;M:蛋氨酸;F:苯丙氨酸;P:脯氨酸;S:丝氨酸;T:苏氨酸;W:色氨酸;Y:酪氨酸;V:缬氨酸
Fig. 1 Amino acid sequence alignment of γ-TMT A: Alanine. R: Arginine. N: Asparagine. D: Aspartic acid. C: Cystine. Q: Glutarnine. E: Lutamic acid. G: Glycine. H: Histidine. I: Isoleucine. L: Leucine. K: Lysine. M: Methionine. F: Phenylalanine. P: Proline. S: Serine. T: Threonine. W: Tryptophan. Y: Tyrosine. V: Valine
图3 激素和逆境胁迫环境下SPAD值 不同小写字母表示与对照相比差异显著(P<0.05)。下同
Fig. 3 SPAD values in hormonal and adversely stress environment Different lowercase letters indicate significant differences compared with the control(P<0.05). The same below
标准品 Standard | 回归方程 Regression equation | R2 | 检出限 Detection limit/(μg·g-1) |
---|---|---|---|
α-生育酚 α-Tocopherol | Y=5.281 3X+0.104 2 | 1 | 0.17 |
γ-生育酚 γ-Tocopherol | Y=22.477X+0.258 3 | 0.999 7 | 0.03 |
δ-生育酚 δ-Tocopherol | Y=25.108X-1.6 | 0.999 8 | 0.03 |
表2 生育酚标准品的回归方程与检出限
Table 2 Regression equation and detection limit of tocopherol standard
标准品 Standard | 回归方程 Regression equation | R2 | 检出限 Detection limit/(μg·g-1) |
---|---|---|---|
α-生育酚 α-Tocopherol | Y=5.281 3X+0.104 2 | 1 | 0.17 |
γ-生育酚 γ-Tocopherol | Y=22.477X+0.258 3 | 0.999 7 | 0.03 |
δ-生育酚 δ-Tocopherol | Y=25.108X-1.6 | 0.999 8 | 0.03 |
不同处理 Different treatment | α-生育酚 α-tocopherol/(μg·g-1) | γ-生育酚 γ-tocopherol/(μg·g-1) | δ-生育酚 δ-tocopherol/(μg·g-1) | 总维生素E Total vitamin E/(μg·g-1) |
---|---|---|---|---|
CK | 21.35±9.41a | 93.04±42.07a | 17.36±7.52a | 131.75±55.16a |
PEG | 12.28±4.81b | 89.35±34.07a | 9.77±5.26b | 111.40±43.08b |
NaCl | 15.26±0.29b | 109.37±15.00b | 10.32±0.44b | 124.25±14.47a |
H2O2 | 8.93±0.61b | 105.83±7.39b | 3.86±0.38b | 118.62±8.26b |
ABA | 11.95±2.14b | 106.33±7.45b | 8.93±0.63b | 127.21±8.77a |
SA | 11.96±3.21b | 81.33±25.48b | 9.22±1.11b | 102.50±25.49b |
表3 激素和逆境胁迫环境下大豆叶中维生素E含量分析
Table 3 Analysis of vitamin E content in soybean leaves under hormonal and adversely stress environment
不同处理 Different treatment | α-生育酚 α-tocopherol/(μg·g-1) | γ-生育酚 γ-tocopherol/(μg·g-1) | δ-生育酚 δ-tocopherol/(μg·g-1) | 总维生素E Total vitamin E/(μg·g-1) |
---|---|---|---|---|
CK | 21.35±9.41a | 93.04±42.07a | 17.36±7.52a | 131.75±55.16a |
PEG | 12.28±4.81b | 89.35±34.07a | 9.77±5.26b | 111.40±43.08b |
NaCl | 15.26±0.29b | 109.37±15.00b | 10.32±0.44b | 124.25±14.47a |
H2O2 | 8.93±0.61b | 105.83±7.39b | 3.86±0.38b | 118.62±8.26b |
ABA | 11.95±2.14b | 106.33±7.45b | 8.93±0.63b | 127.21±8.77a |
SA | 11.96±3.21b | 81.33±25.48b | 9.22±1.11b | 102.50±25.49b |
不同处理 Different treatment | α-生育酚 α-tocopherol/(μg·g-1) | γ-生育酚 γ-tocopherol/(μg·g-1) | δ-生育酚 δ-tocopherol/(μg·g-1) | 总维生素E Total vitamin E/(μg·g-1) |
---|---|---|---|---|
CK | 0.00±0.00 | 8.63±0.69a | 0.57±0.10a | 9.20±0.64a |
PEG | 0.00±0.00 | 3.21±0.47b | 0.68±0.22b | 3.89±0.27b |
NaCl | 0.00±0.00 | 1.00±0.27b | 0.69±0.11b | 1.68±0.29b |
表4 激素和逆境胁迫环境下大豆茎中维生素E含量分析
Table 4 Analysis of vitamin E content in soybean stems under hormonal and adversely stress environment
不同处理 Different treatment | α-生育酚 α-tocopherol/(μg·g-1) | γ-生育酚 γ-tocopherol/(μg·g-1) | δ-生育酚 δ-tocopherol/(μg·g-1) | 总维生素E Total vitamin E/(μg·g-1) |
---|---|---|---|---|
CK | 0.00±0.00 | 8.63±0.69a | 0.57±0.10a | 9.20±0.64a |
PEG | 0.00±0.00 | 3.21±0.47b | 0.68±0.22b | 3.89±0.27b |
NaCl | 0.00±0.00 | 1.00±0.27b | 0.69±0.11b | 1.68±0.29b |
图5 激素及逆境胁迫环境下γ-TMT1的表达分析 A、D:SA处理下叶、根中相对表达水平;B、E:ABA处理下叶、根中相对表达水平;C、F:H2O2处理下叶、根中相对表达水平;G、I:PEG处理下叶、根中相对表达水平;H、J:NaCl处理下叶、根中相对表达水平。下同
Fig. 5 Analysis of γ-TMT1 gene expressions in hormonal and advsrsely stress environment A, D: Relative expression in the leaf and root under SA treated. B, E: Relative expression in the leaf and root under ABA treated. C, F: Relative expression in the leaf and root under H2O2 treated. G, I: Relative expression in the leaf and root under PEG treated. H, J: Relative expression in the leaf and root under NaCl treated. The same below
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