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

A technology of water yellow skin and stress resistance, which is applied in the fields of application, genetic engineering, plant gene improvement, etc., and can solve the problem of not being cloned from water yellow skin

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

AI Technical Summary

Problems solved by technology

Many genes of unknown function have been cloned from different leguminous plants such as alfalfa, chickpea, holly and soybean, but no genes related to abiotic stress have been cloned from P.

Method used

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

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Screening of the gene encoding the stress tolerance-related protein MpSRG of Pseudomonas chinensis and its full-length cDNA clone

[0039] 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 ...

Embodiment 2

[0043] Example 2 The expression characteristics of MpSRG gene of Pygnus chinensis under the treatment of salt stress

[0044] 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 MpSRG gene under salt stress, and the primers were as follows:

[0045]

[0046] The result is as figure 1 As shown, the expression of MpSRG gene was obviously induced by NaCl stress. After salt stress, the expression level of MpSRG gene increased in both roots and leaves, and did not decreas...

Embodiment 3

[0047] Example 3 Construction of MpSRG Recombinant Plant Expression Vector PBI121-MpSRG

[0048] For the construction process of MpSRG recombinant plant expression vector PBI121-MpSRG, please refer to figure 2 .

[0049] Using the cDNA obtained by reverse transcription of the total RNA of Pumice chinensis as a template, use specific primers containing BamHI and SacI linker sequences to PCR amplify MpSRG (sequence 1 in the sequence listing); then BamHI and SacI double-enzyme digest the PCR product and plasmid pBI121 , recovered, ligated with T4DNA ligase, and inserted MpSRG 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-MpSRG.

[0050] The specific primer sequences containing BamHI and SacI linker sequences are as follows:

[0051] 5'-CCGGGATCCATGCTTCACACAATCAAAACG-3'

[0052] 5'-CACGAGCTCTTAAGCATTAGCGTTGGTCC-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 MpSRG, and the base sequence of the pongamia pinnata stress tolerance relative gene MpSRG is shown as SEQ ID NO: 1. An MpSRG recombination plant expression carrier is constructed, arabidopsis is converted by agrobacterium, and MpSRG gene-converted arabidopsis is obtained. The result shows that the MpSRG over expresses in the arabidopsis, the salinity tolerance of the transgenic arabidopsis is improved, and the MpSRG 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-resistant related gene MpSRG and its coded 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 cope with these adverse factors in the environment through various ways. With the development of sequencing technology, especially the widespread application of transcriptome sequencing technology, many genes with unknown functions have been found in plants and may be involved in the regulation of plant stress tolerance. The functions of these new genes are unknown, and some of the...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/82C12N5/10C07K14/415A01H5/00
Inventor 郑易之黄健子黄荣峰陈受宜张万科严昊
Owner SHENZHEN UNIV
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