Functional Characterization of a Zinc-finger-like Gene from Populus trichocarpa

ZHAO Xue-cai; ZHENG Tang-chun; ZANG Li-na; QU Guan-zheng

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Functional Characterization of a Zinc-finger-like Gene from Populus trichocarpa

1.
State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, Heilongjiang, China

Received Date: 2012-10-10

Abstract

In the present study, a zinc-finger-like cDNA (ZFL) was isolated from the cDNA of Populus trichocarpa leaves, named PtrZFL. The open reading frame (ORF) of the gene is 315 bp in length and encodes a predicated polypeptide of 104 amino acids with a molecular weight of about 11.202 kDa, and the theoretical isoelectric point of which is 9.83. The expression of PtrZFL in P. trichocarpa in response to abiotic stress was studied by QRT-PCR, the results showed that the PtrZFL was response to treatments of mannitol, NaCl, H2O2, ABA and cold (4℃). The PtrZFL expression level was the highest 3 hours after treated by ABA and then decreased. The PtrZFL was rapidly and highly induced at the 6th hour when exposed to low temperature of 4℃ and kept high expression level with the continuing treatment. A 1000 bp sequence upstream of PtrZFL gene was cloned from the genomic DNA of P. trichocarpa by PCR based on the poplar genome sequence. Sequence analysis showed that the promoter of the PtrZFL gene contained stress-induced cis-acting elements (including cold, dehydration, frozen and ABA responsive elements ABRE, MYB and WRKY). The GUS activity was almost detected in whole plant of transgenic Arabidopsis, suggesting that this promoter could control the gene expression in the whole plant. Moreover, strong activity was detected in roots and mature leaves, and only weak expression could be detected in other organs.
- poplar,
- PtrZFL,
- QRT-PCR,
- promoter,
- stress

References

[1]	黄骥, 张红生. TFⅢA型锌指蛋白及在提高植物耐逆性中的作用[J]. 遗传, 2007, 29 (8) : 915-922
[2]	黄骥, 王建飞, 张红生. 植物C2H2型锌指蛋白结构和功能[J]. 遗传, 2004, 26 (3) : 414-418
[3]	Seong E S, Wang M H. A novel CaAbsi1 gene induced by early-abiotic stresses in pepper[J]. Biochemistry and Molecular Biology, 2008, 41 (1) : 86-91
[4]	Liu Q L, Xu K D, Ma N, et al. Isolation and functional characterization of DgZFP: a gene encoding a Cys2/His2-type zinc finger protein in chrysanthemum[J]. Japanese Journal of Applied Physics, 2010, 37 (2) :1137-1142
[5]	Sakamoto H, Maruyama K, Sakuma Y, et al. Arabidopsis Cys2/His2-Type Zinc-Finger Proteins Function as Transcription Repressors under Drought, Cold, and High-Salinity Stress Conditions[J]. Plant Physiology, 2004, 136 (1) : 2734-2746
[6]	Yan An, Yucheng Wang, Lingling Lou, et al. A novel Zinc-finger-like gene from Tamarix hispida is involved in salt and osmotic tolerance[J]. Journal of Plant Research, 2011, 124 (6): 689-697
[7]	曲冠证, 郑唐春, 邵龙婷, 等. 烟草类锌指基因NtZFL基因的功能分析[J]. 植物研究, 2012, 32 (4): 430-436
[8]	曲冠证, 郑唐春,杜兆伟, 等. 柽柳类锌指基因ThZFL的功能分析[J]. 东北林业大学学报, 2012,40 (7):6-10,70
[9]	卢孟柱, 胡建军. 我国转基因杨树的研究及应用现状[J] . 林业科技开发, 2006, 20 (6) : 1- 4
[10]	张红梅, 夏新莉, 尹伟伦. 毛果杨的组织培养与快速繁殖[J]. 植物生理学通讯, 2009, 45 (1): 53
[11]	Tamura K, Dudley J, Nei M, et al. MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0[J]. Mol Biol Evol, 2007, 24 (8) :1596-1599
[12]	Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2^-ΔΔCT method[J]. Methods, 2001, 25: 402 - 408
[13]	Doyle J J, Doyle J L. A rapid DNA isolation procedure for small quantities of fresh leaf tissue[J]. Phytochem Bull, 1987, 19: 11-15
[14]	Higo K, Ugawa Y, Iwamoto M, et al. Plant cis-acting regulatory DNA elements (PLACE) database[J]. Nucleic Acids Research ,1999, 27 (1) : 297-300
[15]	Prestridge D S. SIGNAL SCAN:A computer program that scans DNA sequences for eukaryotic transcriptional elements[J]. CABIOS, 1991, 7 (2) : 203-206
[16]	Lescot M, Dehais P, Moreau Y, et al. PlantCARE: a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences[J]. Nucleic Acids Res Database Issue, 2002, 30 (1) : 325-327
[17]	Jefferson R A, Kavanagh T A, Bevan M W. GUS fusions:β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants[J]. EMBO J, 1987,6 (13) : 3901-3907
[18]	Takatsuji H. Zinc-finger transcription factors in plants[J]. Cell Mol Life Sci, 1998, 54 (6) : 582-596
[19]	郑唐春, 施晓文, 臧丽娜, 等. 拟南芥类锌指基因AT3G02790/AT5G16470纯合双突变的筛选及其表型分析[J]. 东北林业大学学报, 2012, 40 (9) : 101-106
[20]	郑唐春, 臧丽娜, 曲冠证, 等.柽柳类锌指基因ThZFL酵母诱饵表达载体的构建及其表达验证[J]. 东北林业大学学报, 2011,39 (12) : 4-8,15
[21]	Gilmour S J, Artus N N, Thomashow M F. cDNA sequence analysis and expression of two cold-regulated genes of Arabidopsis thaliana[J]. Plant Mol Biol, 1992, 18 (1) : 13-21
[22]	郭晋艳, 郑晓瑜, 邹翠霞, 等. 植物非生物胁迫诱导启动子顺式元件及转录因子研究进展[J]. 生物技术通报, 2011, 28 (4) : 16-20,30
[23]	林元震, 张志毅, 林善枝, 等. 运用基因组和EST 数据库进行电子克隆分离杨树功能基因的策略[J]. 分子植物育种, 2007, 5 (4) : 583-587

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

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Functional Characterization of a Zinc-finger-like Gene from Populus trichocarpa

1. State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, Heilongjiang, China

Received Date: 2012-10-10

Abstract: In the present study, a zinc-finger-like cDNA (ZFL) was isolated from the cDNA of Populus trichocarpa leaves, named PtrZFL. The open reading frame (ORF) of the gene is 315 bp in length and encodes a predicated polypeptide of 104 amino acids with a molecular weight of about 11.202 kDa, and the theoretical isoelectric point of which is 9.83. The expression of PtrZFL in P. trichocarpa in response to abiotic stress was studied by QRT-PCR, the results showed that the PtrZFL was response to treatments of mannitol, NaCl, H2O2, ABA and cold (4℃). The PtrZFL expression level was the highest 3 hours after treated by ABA and then decreased. The PtrZFL was rapidly and highly induced at the 6th hour when exposed to low temperature of 4℃ and kept high expression level with the continuing treatment. A 1000 bp sequence upstream of PtrZFL gene was cloned from the genomic DNA of P. trichocarpa by PCR based on the poplar genome sequence. Sequence analysis showed that the promoter of the PtrZFL gene contained stress-induced cis-acting elements (including cold, dehydration, frozen and ABA responsive elements ABRE, MYB and WRKY). The GUS activity was almost detected in whole plant of transgenic Arabidopsis, suggesting that this promoter could control the gene expression in the whole plant. Moreover, strong activity was detected in roots and mature leaves, and only weak expression could be detected in other organs.

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Reference (23)

[1]	黄骥, 张红生. TFⅢA型锌指蛋白及在提高植物耐逆性中的作用[J]. 遗传, 2007, 29 (8) : 915-922
[2]	黄骥, 王建飞, 张红生. 植物C2H2型锌指蛋白结构和功能[J]. 遗传, 2004, 26 (3) : 414-418
[3]	Seong E S, Wang M H. A novel CaAbsi1 gene induced by early-abiotic stresses in pepper[J]. Biochemistry and Molecular Biology, 2008, 41 (1) : 86-91
[4]	Liu Q L, Xu K D, Ma N, et al. Isolation and functional characterization of DgZFP: a gene encoding a Cys2/His2-type zinc finger protein in chrysanthemum[J]. Japanese Journal of Applied Physics, 2010, 37 (2) :1137-1142
[5]	Sakamoto H, Maruyama K, Sakuma Y, et al. Arabidopsis Cys2/His2-Type Zinc-Finger Proteins Function as Transcription Repressors under Drought, Cold, and High-Salinity Stress Conditions[J]. Plant Physiology, 2004, 136 (1) : 2734-2746
[6]	Yan An, Yucheng Wang, Lingling Lou, et al. A novel Zinc-finger-like gene from Tamarix hispida is involved in salt and osmotic tolerance[J]. Journal of Plant Research, 2011, 124 (6): 689-697
[7]	曲冠证, 郑唐春, 邵龙婷, 等. 烟草类锌指基因NtZFL基因的功能分析[J]. 植物研究, 2012, 32 (4): 430-436
[8]	曲冠证, 郑唐春,杜兆伟, 等. 柽柳类锌指基因ThZFL的功能分析[J]. 东北林业大学学报, 2012,40 (7):6-10,70
[9]	卢孟柱, 胡建军. 我国转基因杨树的研究及应用现状[J] . 林业科技开发, 2006, 20 (6) : 1- 4
[10]	张红梅, 夏新莉, 尹伟伦. 毛果杨的组织培养与快速繁殖[J]. 植物生理学通讯, 2009, 45 (1): 53
[11]	Tamura K, Dudley J, Nei M, et al. MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0[J]. Mol Biol Evol, 2007, 24 (8) :1596-1599
[12]	Livak K J, Schmittgen T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2^-ΔΔCT method[J]. Methods, 2001, 25: 402 - 408
[13]	Doyle J J, Doyle J L. A rapid DNA isolation procedure for small quantities of fresh leaf tissue[J]. Phytochem Bull, 1987, 19: 11-15
[14]	Higo K, Ugawa Y, Iwamoto M, et al. Plant cis-acting regulatory DNA elements (PLACE) database[J]. Nucleic Acids Research ,1999, 27 (1) : 297-300
[15]	Prestridge D S. SIGNAL SCAN:A computer program that scans DNA sequences for eukaryotic transcriptional elements[J]. CABIOS, 1991, 7 (2) : 203-206
[16]	Lescot M, Dehais P, Moreau Y, et al. PlantCARE: a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences[J]. Nucleic Acids Res Database Issue, 2002, 30 (1) : 325-327
[17]	Jefferson R A, Kavanagh T A, Bevan M W. GUS fusions:β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants[J]. EMBO J, 1987,6 (13) : 3901-3907
[18]	Takatsuji H. Zinc-finger transcription factors in plants[J]. Cell Mol Life Sci, 1998, 54 (6) : 582-596
[19]	郑唐春, 施晓文, 臧丽娜, 等. 拟南芥类锌指基因AT3G02790/AT5G16470纯合双突变的筛选及其表型分析[J]. 东北林业大学学报, 2012, 40 (9) : 101-106
[20]	郑唐春, 臧丽娜, 曲冠证, 等.柽柳类锌指基因ThZFL酵母诱饵表达载体的构建及其表达验证[J]. 东北林业大学学报, 2011,39 (12) : 4-8,15
[21]	Gilmour S J, Artus N N, Thomashow M F. cDNA sequence analysis and expression of two cold-regulated genes of Arabidopsis thaliana[J]. Plant Mol Biol, 1992, 18 (1) : 13-21
[22]	郭晋艳, 郑晓瑜, 邹翠霞, 等. 植物非生物胁迫诱导启动子顺式元件及转录因子研究进展[J]. 生物技术通报, 2011, 28 (4) : 16-20,30
[23]	林元震, 张志毅, 林善枝, 等. 运用基因组和EST 数据库进行电子克隆分离杨树功能基因的策略[J]. 分子植物育种, 2007, 5 (4) : 583-587

Functional Characterization of a Zinc-finger-like Gene from Populus trichocarpa

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通讯作者: 陈斌, bchen63@163.com

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Functional Characterization of a Zinc-finger-like Gene from Populus trichocarpa

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