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Picea wilsonii transcription factor PwHAP5 and coding gene and application thereof

A technology for encoding gene and transgenic plant, which is applied to the transcription factor PwHAP5 of P. chinensis and its encoding gene and application field

Inactive Publication Date: 2012-05-30
BEIJING FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have shown that HAP transcription factors can participate in various growth and physiological and biochemical processes of plants, but there are no reports on their resistance to salt and drought in the process of seed germination

Method used

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  • Picea wilsonii transcription factor PwHAP5 and coding gene and application thereof
  • Picea wilsonii transcription factor PwHAP5 and coding gene and application thereof
  • Picea wilsonii transcription factor PwHAP5 and coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1, Discovery of transcription factor PwHAP5 and its coding gene

[0043] 1. Processing of green stem pollen

[0044] On the sunny morning when the inflorescences of Qinggan (Zhang LY, Lin JX, et a1.Plant Science, 2007, 172: 1210-1217.) (Piceawilsonii Mast., collected from the Institute of Botany, Chinese Academy of Sciences, Beijing) were open, use sterilized forceps to take Put the yellow pollen pieces on white clean paper, put them into a sterilized dry glass bottle, dry them in the shade indoors, then pack them into dry glass bottles, seal them, and store them in a -20°C refrigerator spare. The pollen stored in the -20°C refrigerator was transferred to 4°C for recovery for 12 hours, and then transferred to room temperature for 2 hours of recovery. The revived pollen was placed in a liquid medium and germinated at room temperature (25±1°C). The composition of the medium includes: 12% sucrose, 0.03% calcium nitrate or 0.1% calcium nitrate, 0.01% boric acid, ...

Embodiment 2

[0051] Example 2, analysis of tissue expression of PwHAP5 gene by semi-quantitative RT-PCR

[0052] Extract the RNA from the pollen, needle leaf, bark and root tissue of the green stem, and reverse transcribe to synthesize cDNA; The primers were used for PCR amplification to detect the expression of PwHAP5 gene in different tissues of A. sinensis. At the same time, the EF1-α gene was used as an internal reference, and the 5' primer for amplifying the EF1-α gene was: 5'-AACTGGAGAAGGAACCCAAG-3', and the 3' primer was: 5'-AACGACCCAATGGAGGATAC-3'.

[0053] PCR reaction conditions: pre-denaturation at 94°C for 5 minutes; then 10 sec at 94°C, 10 sec at 56°C, 20 sec at 72°C, a total of 28 cycles; extension at 72°C for 5 min.

[0054] The expression of PwHAP5 gene in each tissue is as follows: figure 1 shown. The results showed that PwHAP5 was expressed in all tissues, the expression level was lower in roots and the highest in needles.

Embodiment 3

[0055] Embodiment 3, the acquisition of transgenic Arabidopsis and its detection

[0056] 1. Construction of recombinant expression vector

[0057] 1. Acquisition of PwHAP5 gene

[0058] Prepare the following primer pairs:

[0059] Primer 1: 5'-ATGGATCAGCAGCAGCCCA-3';

[0060] Primer 2: 5'-TCAACTGCTGCCATGAGGAGG-3'.

[0061] With Qinggan (Picea wilsonii Mast.,) (Zhang LY, Lin JX, et al.Plant Science, 2007, 172: 1210-1217.) (the non-patent literature that has recorded this material is Zhang LY, Hao HQ, Lin JX .The localization of Rac GTPase in Picea willsonii pollen tubes implies roles in tube growth and the movement of the tube nucleus and sperm cells.Plant Science, 2007, 172:1210-1217, the public can obtain from Beijing Forestry University) cDNA as a template, using The above primers 1 and 2 were amplified by PCR. Sequencing showed that the nucleotide sequence of the amplified product was shown in sequence 1 in the sequence listing. There were 606 nucleotides in sequence 1...

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Abstract

The invention discloses a picea wilsonii transcription factor PwHAP5 and a coding gene and application thereof. The PwHAP5 is protein disclosed in 1) or 2): 1) the protein composed of an amino acid sequence disclosed in a sequence 2 in a sequence table; 2) the protein which is performed with substitution and / or deletion and / or adding of one of multiple amino acid residue by the amino acid sequence disclosed in the sequence 2 in the sequence table, is relevant to plant seed germination and is derived by 1). After the coding gene of PwHAP5 protein is guided into arabidopsis, the germination rate of the seed of transgenosis arabidopsis is obviously higher than that of wild arabidopsis in high-salt and wild environment.

Description

technical field [0001] The invention relates to the transcription factor PwHAP5 and its coding gene and application. Background technique [0002] The production of crops is seriously affected by abiotic stresses such as salinity and drought. About 20% of the cultivated land and 50% of the irrigated fields in the world are threatened by different degrees of salt damage, and there is an increasing trend year by year. The area of ​​saline land in my country is about 140 million mu, accounting for nearly 7% of the total cultivated land area. In addition, there are more than 200 million mu of saline wasteland. The total area of ​​arid and semi-arid regions in the world accounts for 34.9% of the total land area; the area of ​​arid and semi-arid cultivated land in my country accounts for 51% of the total area. It is an effective way to improve the salt tolerance and drought resistance of crops through genetic modification. Therefore, it is of great theoretical and practical sig...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21A01H5/00
Inventor 张凌云于彦丽李彦泽郑成超
Owner BEIJING FORESTRY UNIVERSITY
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