番茄<i>SlNAC1</i>基因启动子低温、高温和ABA应答顺式元件分析<sup>*</sup>

doi:10.13523/j.cb.2305017

中国生物工程杂志

2023, Vol. 43

Issue (11): 16-26 DOI: 10.13523/j.cb.2305017

研究报告

番茄SlNAC1基因启动子低温、高温和ABA应答顺式元件分析^*

李德鑫^1,²,乐露³,陶向⁴,廖海¹,周嘉裕^1,^**(

),黄维藻^2,^**(

)

1 西南交通大学生命科学与工程学院成都 610031
2 中国科学院成都生物研究所成都 610041
3 阿坝师范学院资源与环境学院成都 623002
4 四川师范大学生命科学学院成都 610101

Analysis of Cold-, Heat-, and ABA-responsive cis Elements in Tomato SlNAC1 Promoter

LI De-xin^1,²,YUE Lu³,TAO Xiang⁴,LIAO Hai¹,ZHOU Jia-yu^1,^**(

),HUANG Wei-zao^2,^**(

)

1 School of Life Science and Engineering, Southwest Jiaotong University, Chengdu 610031, China
2 Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu 610041, China
3 College of Resources and Environment, Aba Teachers University, Chengdu 623002, China
4 College of Life Science, Sichuan Normal University, Chengdu 610101, China

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摘要：

番茄转录因子SlNAC1调控多种生物和非生物胁迫应答,但其上游转录调控因子至今不明,限制了对其胁迫应答机制的理解。构建了系列5'-缺失的SlNAC1启动子(起始密码子上游2 039 bp、1 508 bp、1 373 bp和777 bp)驱动的GUS转基因烟草,并定量分析了其在低温、高温和ABA诱导下的GUS酶活,以鉴定SlNAC1启动子中的低温、高温和ABA应答顺式元件。结果显示,低温和高温诱导后,2 039 bp启动子转基因烟草GUS酶活的增长显著高于其他转基因烟草和野生型烟草;而ABA诱导后,1 508 bp启动子转基因烟草GUS酶活的增长显著高于其他转基因烟草和野生型烟草。这些结果说明低温和高温应答顺式元件位于-2 039~-1 508 bp区间,而ABA应答顺式元件位于-1 508~-1 373 bp区间。启动子顺式元件预测分析显示,-2 039 bp~-1 508 bp区间只有1个低温/高温/干旱/盐应答顺式元件DRE/CRT,而-1 508 bp~-1 373 bp区间只有1个ABA应答顺式元件ABRE。因此,这两个顺式元件将作为候选元件用于后续SlNAC1上游调控转录因子的筛选。

关键词： 转录因子; SlNAC1; 启动子; 顺式元件; 胁迫应答

Abstract:

Tomato transcription factor SlNAC1 has been reported to regulate several biotic and abiotic stress responses, but its upstream regulatory transcription factors remain unknown, which limits our understanding of its molecular mechanism of stress responses. We constructed series of 5'-deleted SlNAC1 promoters (2 039 bp, 1 508 bp, 1 373 bp and 777 bp upstream of start codon) -driven GUS transgenic Nicotiana benthamiana and analyzed quantitatively their GUS activity under cold, heat and ABA treatment conditions in order to identify cold-, heat-, and ABA-responsive cis elements. Our results show that the GUS activity of 2 039 bp promoter-driven transgenic Nicotiana benthamiana increased much higher than that of any other transgenic Nicotiana benthamiana and wild-type Nicotiana benthamiana after cold and heat treatment, while the GUS activity of 1 508 bp promoter-driven transgenic Nicotiana benthamiana increased much higher than that of any other transgenic Nicotiana benthamiana and wild-type Nicotiana benthamiana after ABA treatment. These results indicate that both cold-responsive and heat-responsive cis elements were located in the region from -2 039 bp to -1 508 bp, and ABA-responsive cis element(s) was(were) located in the region from -1 508 bp to -777 bp. According to cis elements' prediction of SlNAC1 promoter, there were only one cold/heat/drought/salt-responsive cis element DRE/CRT in the region from -2 039 bp to -1 508 bp and only one ABA-responsive cis element ABRE in the region from -1 508 bp to -777 bp. Therefore, these two cis elements will be used as candidates for subsequent site-directed mutagenesis validation and screening of the upstream regulatory transcription factor of SlNAC1.

Key words: Transcription factor SlNAC1 Promoter cis element Stress-responsive

收稿日期: 2023-05-11 出版日期: 2023-12-01

ZTFLH:

Q789

基金资助: *四川省国际科技创新合作(2020YFH0003)

通讯作者: **周嘉裕,黄维藻 E-mail: spinezhou@home.swjtu.edu.cn;huangwz@cib.ac.cn

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引用本文:

李德鑫, 乐露, 陶向, 廖海, 周嘉裕, 黄维藻. 番茄SlNAC1基因启动子低温、高温和ABA应答顺式元件分析^*[J]. 中国生物工程杂志, 2023, 43(11): 16-26.

LI De-xin, YUE Lu, TAO Xiang, LIAO Hai, ZHOU Jia-yu, HUANG Wei-zao. Analysis of Cold-, Heat-, and ABA-responsive cis Elements in Tomato SlNAC1 Promoter. China Biotechnology, 2023, 43(11): 16-26.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.2305017 或 https://manu60.magtech.com.cn/biotech/CN/Y2023/V43/I11/16

表1 qRT-PCR引物信息

表2 引物信息

图1 低温、高温和ABA处理下SlNAC1的相对表达水平

图2 SlNAC1启动子胁迫应答顺式元件预测

图3 系列5'-缺失SlNAC1启动子驱动的GUS表达载体及其T-DNA在染色体上的插入位点示意图

图4 低温处理下GUS染色和GUS酶活定量

图5 高温处理下GUS染色和GUS酶活定量

图6 ABA处理下GUS染色和GUS酶活定量

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