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Application of GhTZF1 gene to reinforcement on drought resistance of plants and senility delay

A gene and drought tolerance technology is applied in the field of plant genetic engineering to achieve the effect of improving tolerance and improving drought tolerance.

Inactive Publication Date: 2013-12-25
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report that TZF genes isolated and cloned from cotton are involved in plant drought resistance and delaying drought-induced senescence

Method used

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  • Application of GhTZF1 gene to reinforcement on drought resistance of plants and senility delay
  • Application of GhTZF1 gene to reinforcement on drought resistance of plants and senility delay
  • Application of GhTZF1 gene to reinforcement on drought resistance of plants and senility delay

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Isolation, cloning and expression pattern analysis of GhTZF1 gene

[0031] The applicant established the cell wall regeneration SSH library (Yang et al., 2008, Expression profile analysis of genes involved in cell wall regeneration during protoplast culture in cotton by suppression) in the preliminary work of the State Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University Subtractive hybridization and macroarray.J Exp Bot,59:3661-3674) isolated an EST sequence (NCBI EST accession number FF403655, submitted in April 2009), the EST sequence was compared in NCBI GenBank by tBLASTx Yes, it was found that this sequence may be a CCCH-type transcription factor. This EST sequence does not contain a complete open reading frame. We used the RACE (rapid-amplification of cDNA ends) method to perform rapid cloning of cDNA ends by PCR to obtain the complete coding sequence of this gene. The specific steps are as follows: A. RNA extraction and cDNA a...

Embodiment 2

[0037] Embodiment 2: Construction of GhTZF1 gene plant overexpression vector

[0038]Primers were designed according to the obtained sequence of SEQ ID NO: 1 for the construction of expression vectors, and the linker bases for BP-LR reaction were added to both ends of the primers respectively. and GhTZF1BP-R (5'-GGGGACCACTTTGTACAAGAAA GCTGGGTTTCATTTTCACCAACTCAGATAC-3'), using the T-GhTZF1 plasmid as a template for PCR amplification, the PCR reaction conditions were: 94°C pre-denaturation for 3min; 94°C for 30sec, 58°C for 30sec, 72°C for 1min, A total of 28 cycles; 72°C extension for 7 minutes, PCR amplification to obtain a PCR product containing the complete ORF. The PCR product was ligated into pDONR by BP reaction TM 221 carrier (purchased from Invitrogen, the United States, see the effect on Figure 3A , after transforming Escherichia coli competent cells TOP10, use GhTZF1-specific primers GhTZF1BP-F (5'-GGGGACAAGTTTGTACAAAAAGCAGGCTTCCAAAAATGATG ATCGGAGA-3') and GhTZF1BP...

Embodiment 3

[0041] Genetic transformation and screening identification of embodiment 3GhTZF1 gene

[0042] A. Preparation of Arabidopsis

[0043] The test material is wild-type Arabidopsis thaliana L.Columbia ecotype. Wild-type Arabidopsis thaliana seeds were sown with special nutrient soil for Arabidopsis planting after vernalization (Pei Lei brand, produced in Zhenjiang, Jiangsu, China, commercial nutrient soil) and placed in an artificial cultivation room (16 hours of light, 22±2°C ) and wait for the Arabidopsis to grow to about 4 leaves to control the growth density of Arabidopsis. The Arabidopsis can be transformed when it begins to flower about 6 weeks after its growth, and the Arabidopsis is watered enough the day before the transformation.

[0044] B. Activation of Agrobacterium

[0045] Take out the glycerol tube of the GV3101 strain containing the target gene (that is, the cloned GhTZF1 gene of the present invention) from the ultra-low temperature refrigerator and melt it on ...

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Abstract

The invention belongs to the technical field of plant genetic engineering and particularly relates to isolation and cloning, functional verification and application of a GhTZF1 gene separated from cotton. A cDAN (complementary Deoxyribonucleic acid) sequence of the cloned GhTZF1 disclosed by the invention is shown as SEQ ID NO:1; and an amino acid sequence of a coding gene of the GhTZF1 is shown as SEQ ID NO:2. The invention also discloses application of the gene to improvement on drought tolerance of arabidopsis thaliana and delay on plant senescence under the drought stress.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering. It specifically relates to the functional verification and application of a CCCH-type transcription factor GhTZF1 isolated and identified from cotton. Functional verification showed that the GhTZF1 gene can improve Arabidopsis tolerance to drought and delay drought-induced senescence, and also delay H 2 o 2 induced senescence. The GhTZF1 gene cloned by the invention can be applied to enhance the tolerance of plants to drought and delay the senescence caused by drought through genetic transformation. Background technique [0002] Drought refers to a state in which there is a shortage of adequately available water that results in a reduction in yield, or a state in which there is no rainfall or irrigation for a period of time that affects crop growth (Bernier et al., 2008, Breeding upland rice for drought resistance. Journal of the Science of Food and Agriculture, 88:927-939)....

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/00
Inventor 张献龙杨细燕周婷王丽晨
Owner HUAZHONG AGRI UNIV
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