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Acta Agronomica Sinica ›› 2019, Vol. 45 ›› Issue (3): 469-476.doi: 10.3724/SP.J.1006.2019.84082

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Expression and promoter activity of GhTFL1a and GhTFL1c in Upland cotton

Xiao-Hong ZHANG1,Gen-Hai HU1,Han-Tao WANG2,Cong-Cong WANG2,Heng-Ling WEI2,Yuan-Zhi FU1,Shu-Xun YU2,*()   

  1. 1 College of Life Science and Technology, Henan Institute of Science and Technology / Henan Collaborative Innovation Center of Modern Biological Breeding, Xinxiang 453003, Henan, China
    2 Institute of Cotton Research, Chinese Academy of Agricultural Sciences / State Key Laboratory of Cotton Biology, Anyang 455000, Henan, China
  • Received:2018-06-20 Accepted:2018-12-24 Online:2019-03-12 Published:2019-01-04
  • Contact: Shu-Xun YU E-mail:ysx195311@163.com
  • Supported by:
    This study was supported by the Key Scientific Research Projects of Colleges and Universities in Henan(18A210002);the State Key Laboratory of Cotton Biology Open Fund(CB2018A08)

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Abstract:

In this study, we cloned the phosphatidylethanolamine-binding protein GhTFL1a and GhTFL1c genes from Upland cotton, and analyzed their expression and promoter activity. The results of promoter structure prediction revealed that GhTFL1a promoter contains abscisic acid (ABA) responsiveness elements, drought-induced MYB binding sites and shoot-specific expression and light responsiveness elements, and the promoter region of GhTFL1c contains ethylene-responsive element, drought-induced MYB binding sites and salicylic acid (SA) responsiveness elements. Thus, we constructed the fusion vector pBI121-GhTFL1a-GUS and pBI121-GhTFL1c-GUS, respectively. Transient transformation of tobacco showed that both promoters had the activity to drive the expression of target gene GUS. Quantitative Real-time PCR result indicated that the expression profile of GhTFL1a and GhTFL1c was opposite during different photoperiod treatments of cultivated and semi-wild cotton. Meanwhile, the expression of GhTFL1a was induced by ABA, SA, and salt (NaCl), while GhTFL1c expression was induced by SA, gibberellin (GA) and ABA. Taken together, the results suggest that GhTFL1a and GhTFL1c might be involved in the regulation of response to abiotic stresses (SA and ABA), which could provide a solid foundation for further function identification.

Key words: upland cotton, GhTFL1a, GhTFL1c, expression analysis, promoter activity

Table 1

Primers used in this study"

引物名称
Primer name		 引物序列
Primer sequence (5′-3′)		 用途
Purpose
GhTFL1a-up TCCCTGAGCCACTTACCGTTG 荧光定量PCR qRT-PCR
GhTFL1a-down AAGCGTCTCTCATGTCGTTG 荧光定量PCR qRT-PCR
GhTFL1c-up TCATCTGTTGCCACCAAACCT 荧光定量PCR qRT-PCR
GhTFL1c-down TTCCCTTCCAAACGTGGCATC 荧光定量PCR qRT-PCR
GhACTIN-up ATCCTCCGTCTTGACCTTG 荧光定量PCR qRT-PCR
GhACTIN-down TGTCCGTCAGGCAACTCAT 荧光定量PCR qRT-PCR
GhTFL1a-up ATGTCAAGGGTCCCTGAG 基因克隆 Gene cloning
GhTFL1a-down TTATCTTCTTCTTGCAGCAGTTTC 基因克隆 Gene cloning
GhTFL1c-up ATGGGAGAGCCTCTCATTGTT 基因克隆 Gene cloning
GhTFL1c-down TTAGCGTCTCCTTGCAGCAGT 基因克隆 Gene cloning
pGhTFL1a-up AAGGAATATAGAGCACAACA 启动子克隆 Promoter cloning
pGhTFL1a-down GATGAACAAGACGATGTGTAT 启动子克隆 Promoter cloning
pGhTFL1c-up AGTTTAGATTCTTGTGCGAT 启动子克隆 Promoter cloning
pGhTFL1c-down TCTTGATGACAGTGAATGAA 启动子克隆 Promoter cloning

Fig. 1

Phylogenetic and conserved domain analysis of GhTFL1a and GhTFL1c A: phylogentic analysis of GhTFL1a/c and their homologous genes in Arabidopsis; B: alignment of amino acid sequences of GhTFL1a/c and their homologous genes in Arabidopsis. * indicates the same amino acid, the ID of AtBFT, AtTFL1, and AtATC is NM_125597, U77674, and NM_128315, respectively, the ID of GhTFL1a and GhTFL1c is Gh_A11G0088 and Gh_D04G0971, respectively."

Fig. 2

Expression patterns of GhTFL1a and GhTFL1c genes during photoperiod treatment A and B indicate the expression of GhTFL1a and GhTFL1c in cotton cultivar “CCRI 36”; C and D indicate the expression of GhTFL1a and GhTFL1c in semi-wild cotton. Error bars indicate the standard deviation (±SD)."

Fig. 3

Expression patterns of GhTFL1a and GhTFL1c genes in cultivated cotton and semi-wild race cotton The abscissa shows period, which from left to right is expanded cotyledon, first, second, third, forth, and fifth leaves in long day, and third, forth and fifth leaves in short day."

Table 2

cis-acting elements in promoter of GhTFL1a and GhTFL1c"

基因
Gene		 元件
Element		 序列
Sequence (5′-3′)		 功能
Function
GhTFL1a CE3 GACGCGTGTC ABA和VP1响应元件 ABA and VP1 responsiveness element
MBS TAACTG 干旱诱导的MYB结合位点 MYB binding site involved in drought inducibility
Nodule-site2 CTTAAATTATTTATTT 节结特异因子结合位点 Nodolue specific factor binding site
as-2-box GATAatGATG 顶芽特异表达响应元件 Shoot specific expression element
as-2-box GATAatGATG 顶芽特异表达响应元件 Shoot specific expression element
Circadian CAANNNNATC 昼夜节律控制元件 Circadian rhythms element
GhTFL1c ERE ATTTCAAA 乙烯响应元件 Ethylene responsiveness element
ERE ATTTCAAA 乙烯响应元件 Ethylene responsiveness element
MBS CAACTG 干旱诱导的MYB结合位点 MYB binding site involved in drought inducibility
circadian CAANNNNATC 昼夜节律控制元件 Circadian rhythms element
TCA-element TCAGAAGAGG 水杨酸响应元件 Salicylic acid responsiveness element
TCA-element CAGAAAAGGA 水杨酸响应元件 Salicylic acid responsiveness element

Fig. 4

GhTFL1a expression profiles in the first 24 hours after the roots were treated A: salicylic acid; B: gibberellins; C: abscisic acid; D: NaCl. Error bars on the columns indicate the standard deviation (±SD)."

Fig. 5

GhTFL1c expression profiles in the first 24 hours after the roots were treated A: salicylic acid; B: gibberellins; C: abscisic acid; D: NaCl; Error bars indicate the standard deviation (±SD)."

Fig. 6

Transient expression of GhTFL1a and GhTFL1c promoter in tobacco A: positive control pBI121; B: pGhTFL1a∷GUS; C: pGhTFL1c∷GUS; D: negative control."

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