Crops ›› 2017, Vol. 33 ›› Issue (2): 51-58.doi: 10.16035/j.issn.1001-7283.2017.02.009

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Cloning and Sequence Analysis of GmWRI1a Gene Promoter in Soybean

Yan Li,Yang Qiang,Shao Yupeng,Li Dandan,Wang Zhikun,Li Wenbin   

  1. Northeast Agricultural University/Key Laboratory of Soybean Biology, Chinese Ministry of Education/Key Laboratory of Soybean Biology and Genetics Breeding,Chinese Agriculture Ministry,Harbin 150030,Heilongjiang,China
  • Received:2016-11-21 Revised:2017-02-26 Online:2017-04-15 Published:2018-08-26
  • Contact: Zhikun Wang,Wenbin Li

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Abstract:

According to the Williams 82’s genomic sequences reported in the Phytozome, primers were designed by Primer Premier 5.0. The promoter product sequence of GmWRI1a gene was amplified by PCR method. The recombinant vectors pGM-T-pGmWRI1a and PBI121-pGmWRI1a were constructed. The positive clones were identified by PCR amplification and restriction enzyme digestion. The full length of the GmWRI1a promoter was 1 687bp. The promoter sequence contained the necessary initiation transcription sites, such as TATA-box,CTTA-box and multiple other cis-acting elements: light responsiveness element, gibberellin response element, cis-acting regulatory element related to meristem expression, drought-inducing elements, and others. Cloning and characterization of the GmWRI1a gene promoter will provide basis for the further study of regulation and functional analysis of GmWRI1a gene in soybean.

Key words: Promoter of GmWRI1a gene, Soybean, Sequence analysis, Cis-regulatory element

Fig.1

DNA extract result of Dongnong 47"

Fig.2

PCR results of GmWRI1a gene promoter"

Fig.3

PCR identify of pGM-T-pGmWRI1a recombinant plasmid"

Fig.4

Sequence comparison result of GmWRI1a gene promoter"

Fig.5

GmWRI1a gene promoter sequential analysis The possible cis-acting elements in the sequence is signed by frames; The black is transcriptional start site;start codon ATG is signed by the double underline"

Table 1

GmWRI1a gene promoter cis-form regulatory sequence position and function"

调控元件Regulatory sequence 位置Position 核心序列Core sequence 预测功能Function
ACE
 797（+）
 ACGTGGA
 光应答元件
Cis-acting element involved in light responsiveness
AE-Box

 176（+）,
1278（+）,
655（–）		 AGAAAC

 光响应模块
Part of a module for light response
ATC-motif
 1227（–）
 AGTAATCT
 光应答元件
Part of a conserved DNA module involved in light responsiveness
CATT-motif
 210（+）
 GCATTC
 光响应一部分
Part of a light responsive
GA-motif
 835（–）
 AAAGATGA
 光响应元件
Part of a light responsive element
G-box
 795（+）,
796（–）		 TGACGTGG
CACGTC		 光反应调节元件
Cis-acting regulatory element involved in light responsiveness
调控元件Regulatory sequence 位置Position 核心序列Core sequence 预测功能Function
GT1-motif

 674（+）,
1056（+）,
914（–）		 GGTTAA

 光反应元件
Light responsive element
CTA-box
 1008（+）
 GCCACT
 表达分生组织相关
Cis-acting regulatory element related to meristem expression
P-Box
 896（–）
 CCTTTTG
 赤霉素应答元件
Gibberellin-responsive element
GARE-motif
 603（+）,
1482（+）		 AAACAGA
TCTGTTG		 赤霉素响应元件
Gibberellin-responsive element
CE3
 802（+）
 GACGCGTGT
 ABA和VP1响应元件
Cis-acting element involved in ABA and VP1 responsiveness
MBS
 791（+）
 TAACTG
 MYB参与抗旱诱导结合位点
MYB binding site involved in drought-inducibility
Skn-1_motif
 362（+）,
664（+）		 GTCAT
 胚乳表达所需的调节元件
Cis-acting regulatory element required for endosperm expression
GCN4_motif

 370（+）,
1005（+）,
818（+）		 TGAGTCA

 胚乳表达顺式调控元件
Cis-regulatory element involved in endosperm expression

Fig.6

PCR identify of pBI-pGmWRI1a recombinant plasmid"

Fig.7

Hind Ⅲ、XbaⅠenzyme identify of pBI-pGmWRI1a recombinant plasmid"

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