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Pongamia pinnata stress tolerance relative gene MpZFP as well as coded protein and application thereof

A kind of water yellow bark and stress-resistant technology, which is applied in the fields of application, genetic engineering, plant gene improvement, etc., and can solve problems such as cloning

Inactive Publication Date: 2015-01-07
SHENZHEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Zinc finger protein transcription factors have been cloned from Arabidopsis, soybean, rice and other crops, but such transcription factors related to adversity stress have not been cloned from the non-salt-tolerant non-model plant Pseudomonas

Method used

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  • Pongamia pinnata stress tolerance relative gene MpZFP as well as coded protein and application thereof
  • Pongamia pinnata stress tolerance relative gene MpZFP as well as coded protein and application thereof
  • Pongamia pinnata stress tolerance relative gene MpZFP as well as coded protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Screening of the Stress Tolerance-Related Protein MpZFP Encoding Gene of Pseudomonas chinensis and its Full-length cDNA Cloning

[0049] Use seawater (equivalent to 30‰NaCl) and fresh water to treat the one-month-old plants of P. japonicus, then select the root and leaf samples treated for 2, 4 and 8 hours for RNA extraction, and then mix the RNA at the three time points in equal amounts , four libraries were constructed (namely root freshwater treatment, root seawater treatment, leaf freshwater treatment and leaf seawater treatment) and then performed Illumina sequencing. Finally, the sequencing results were assembled and spliced ​​to obtain more than 100,000 unigene sequences. The gene was selected according to the following three principles: first, the expression of the gene was significantly up-regulated after salt stress; second, the sequence with the highest homology of the gene came from legume species; third, the genes with the highest similarity to th...

Embodiment 2

[0054] The expression characteristics of the MpZFP gene of Pygnus chinensis under the treatment of embodiment 2 salt stress

[0055] Divide 20 one-month-old seedlings with uniform growth into 5 groups (4 plants in each group) on average, and pour 30‰ NaCl solution to these five groups of seedlings for 0, 2, 4, 8, and 12 hours respectively. The top young leaves and roots of each group of plants were quickly taken and stored at -80°C after quick freezing in liquid nitrogen, and the roots and leaves were weighed to the same weight. Total RNA was extracted by Trizol method and reverse transcribed into cDNA. Real-time fluorescent quantitative PCR was carried out to analyze the expression of the MpZFP gene under salt stress, and the primers used were as follows:

[0056]

[0057] The result is as figure 1 As shown, the expression of MpZFP gene was obviously induced by NaCl stress. After salt stress, the change of MpZFP gene expression in roots was greater than that in leaves, ...

Embodiment 3

[0058] Example 3 Construction of MpZFP recombinant plant expression vector PBI121-MpZFP

[0059] For the construction process of MpZFP recombinant plant expression vector PBI121-MpZFP, please refer to figure 2 .

[0060] Using the cDNA obtained by reverse transcription of the total RNA of Pseudomonas chinensis as a template, use specific primers containing BamHI and SacI linker sequences to PCR amplify MpZFP (SEQ ID NO: 1 in the sequence listing); then BamHI and SacI double-digest the PCR product and plasmid pBI121, recovered, connected with T4DNA ligase, and inserting MpZFP forward into the CaMV35S promoter of the plant expression vector PBI121 between the BamHI and SacI restriction sites to obtain the recombinant plant expression vector PBI121-MpZFP.

[0061] Specific primers containing BamHI and SacI linker sequences are as follows:

[0062] 5'-CCGGGATCCATGAAGAGAGAAAGGGAAGGT-3'

[0063] 5'-CACGAGCTCTCAATTGAAACAATGAACCAAAG-3'.

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Abstract

The invention relates to the field of plant gene engineering and provides a pongamia pinnata stress tolerance relative gene MpZFP, and the base sequence of the pongamia pinnata stress tolerance relative gene MpZFP is shown as SEQ ID NO: 1. An MpZFP recombination plant expression carrier is constructed, arabidopsis is converted by agrobacterium, and MpZFP gene-converted arabidopsis is obtained. The result shows that the MpZFP over expresses in the arabidopsis, the salinity tolerance of the transgenic arabidopsis is improved, and the MpZFP gene has an effect on improvement of plant stress tolerance is proved.

Description

technical field [0001] The invention relates to the field of plant genetic engineering, in particular to a stress tolerance-related gene MpZFP of Pseudomonas chinensis and its encoded protein and application. Background technique [0002] Abiotic stresses such as salinity and drought affect the growth and development of crops, and even cause crop death in severe cases. In order to increase crop yield, cultivating stress-tolerant crop varieties is one of the main coping methods. At present, breeding through genetic engineering has become one of the important methods for cultivating stress-tolerant crops. Higher plants can deal with these unfavorable factors in the environment through a variety of ways, and the transcription factors in them have become one of the key points in the study of plant stress tolerance because they can regulate the expression of multiple downstream genes. Many types of transcription factors related to plant stress tolerance have been discovered, su...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82C12N5/10C07K14/415A01H5/00
Inventor 郑易之黄健子陈受宜黄荣峰张万科陆翔
Owner SHENZHEN UNIV
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