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DREB1C from Medicago truncatula enhances freezing tolerance in transgenic M. truncatula and China Rose (Rosa chinensis Jacq.)

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Abstract

We isolated a DREB orthologue, MtDREB1C, from Medicago truncatula. Its deduced protein contains an AP2 domain of 57 amino acids. Yeast one-hybrid assay revealed that MtDREB1C specifically bound to the dehydration-responsive element (DRE) and activated the expression of HIS3 and LacZ reporter genes. In a transcriptional activation assay, coexpression of the MtDREB1C cDNA resulted in much higher (21.2 times) transactivation of the LacZ reporter gene than experiments performed without MtDREB1C. Transformation of Medicago revealed that overexpression of MtDREB1C suppressed shoot growth, and enhanced the freezing tolerance of M. truncatula. The MtDREB1C gene was transformed into China Rose (Rosa chinensis Jacq.) driven by Arabidopsis rd29A promoter. Southern-blot analysis showed that the target gene was integrated into the genome of a surviving transgenic rose plant. Northern-blot analysis illustrated that robust expression of MtDREB1C was only activated under stress conditions, and the expressed MtDREB1C mRNA reached maximum accumulation 10 h following freezing treatment. The performance of the transgenic line under freezing stress was superior to untransformed controls. This transgenic plant continued to grow, flowered under unstressed conditions, and was phenotypically normal. These facts indicate that the MtDREB1C gene, isolated from Medicago truncatula and driven by the Arabidopsis rd29A promoter, enhanced freezing tolerance in transgenic China Rose significantly without any obvious morphological or developmental abnormality.

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Abbreviations

2,4-D:

2,4-Dichlorophenoxy acetic acid

BD:

binding domain

CaMV35S:

Cauliflower mosaic virus 35S promoter

CBF:

C-repeat/DRE binding factor

CRT:

C-repeat

DRE:

Dehydration-responsive element

DREB:

Dehydration-responsive element binding

EST:

Expressed sequence tag

EREBP:

Ethylene-responsive element binding protein

ERF:

Ethylene-responsive-element-binding factor

ICE1:

Inducer of CBF expression 1

MtDREB1C:

Medicago truncatula DREB1C

ONPG:

o-nitrophenyl β-D-galactopyranoside

rd29A:

Responsive drought 29A

SD medium:

Synthetic dextrose medium

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Acknowledgments

The authors gratefully acknowledge laboratory support provided by Professor Xiaofang Luo at Beijing Forestry University and Xingyao Xiong at Hunan Provincial Key Laboratory for Germplasm Innovation and Utilization of Crops. We also thank Dr. Jingsong Zhang for guidance on yeast hybrid assay. This work is supported by China National “948” Program (Grant No. 2005-4-34, 2007-4-02); National High Technology Program (2006AA10Z182); Special Research Fund for Doctor’s Degree Dissertation in Chinese Universities (20060022012); Research Project in Hunan Universities (09C497); National Natural Science Foundation of China (30371148).

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Authors and Affiliations

  1. College of Biological Sciences and Biotechnology, Beijing Forestry University, 100083, Beijing, People’s Republic of China

    Ji-Ren Chen, Jing-Jing Lü, Tian-xiang Wang & Hua-Fang Wang

  2. College of Horticulture and Gardening, Hunan Agricultural University, 410128, Changsha, People’s Republic of China

    Ji-Ren Chen & Xing-Yao Xiong

  3. Hunan Provincial Key Laboratory for Germplasm Innovation and Utilization of Crops, Changsha, China

    Ji-Ren Chen & Xing-Yao Xiong

  4. Hunan Biological and Electromechanical Polytechnic, Changsha, 410127, People’s Republic of China

    Rong Liu

  5. Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 100101, Beijing, People’s Republic of China

    Shou-Yi Chen

  6. Commonwealth Scientific and Industrial Research Organisation (CSIRO) Plant Industry, Canberra, ACT, Australia

    Lan-Bin Guo

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  1. Ji-Ren Chen
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Correspondence to Shou-Yi Chen or Hua-Fang Wang.

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Ji-Ren Chen and Jing-Jing Lü authors contributed equally to this work.

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Chen, JR., Lü, JJ., Liu, R. et al. DREB1C from Medicago truncatula enhances freezing tolerance in transgenic M. truncatula and China Rose (Rosa chinensis Jacq.). Plant Growth Regul 60, 199–211 (2010). https://doi.org/10.1007/s10725-009-9434-4

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