Soybean bHLH transcription factor gene GmFER and encoded protein and application thereof

A technology for transcription factors and transgenic plants, which is applied in the application field of soybean bHLH transcription factor gene GmFER, tolerance, and can solve the problems of reducing transport, and no report discloses the related application of soybean bHLH transcription factor gene, etc.

Inactive Publication Date: 2017-02-15
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Arabidopsis bHLH genes (FIT, AtbHLH38, and AtbHLH39) are involved in plant responses to Cd stress, and constitutively promote the expression of some genes involved in heavy metal Most of the Cd compartmentalized in the root, reducing the translocation to the shoot (see Wu HL, Chen CL, Du J, et al., Co-Overexpression FIT with AtbHLH38 or AtbHLH39 in Arabidopsis-Enhanced Cadmium Tolerance via Increased Cadmium Sequestration in Roots and Improved Iron Homeostasis of Shoots[J],Plant Physiology,2012,158:790-800), but so far there is no report disclosing the related application of soybean bHLH transcription factor gene

Method used

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  • Soybean bHLH transcription factor gene GmFER and encoded protein and application thereof
  • Soybean bHLH transcription factor gene GmFER and encoded protein and application thereof
  • Soybean bHLH transcription factor gene GmFER and encoded protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1: Cloning of soybean bHLH transcription factor gene GmFER.

[0026] Utilize the method that bioinformatics and routine polymerase chain reaction (PCR) combine to clone the dna sequence of soybean GmFER gene, specific method is as follows:

[0027] The root system of soybean variety Williams 82 was used as experimental material, and total RNA was extracted using SV Total RNAlysis Kit (Promega, USA). According to the method provided by TaKaRa Company, using PrimeScript TM Reverse transcriptase synthesizes cDNA, and uses it as template to carry out PCR amplification, a pair of primers P1 (as shown in SEQ ID NO:3, synthesized by Shanghai Bioengineering Company) and P2 (as shown in SEQ ID NO:3) and P2 (as shown in SEQ ID NO:3) used in the PCR amplification process NO:4, synthesized by Shanghai Bioengineering Company) was designed and synthesized according to the known GmFER sequence in NCBI:

[0028] P1: 5'-ATGGATGTTCACGAAGACACACTCA-3';

[0029] P2: 5'-TCAAGCAGGA...

Embodiment 2

[0031] Example 2: Spatiotemporal distribution of GmFER transcripts and the effect of Cd stress on GmFER gene expression.

[0032] Use the method of semi-quantitative PCR to detect the tissue distribution of GmFER in soybean plants, and take the following measures to ensure the reliability of the detection results: Use the amplification grade DNase I to digest and extract a small amount of genomic DNA contamination in the total RNA , in order to detect whether the genomic DNA is eliminated, a part of the total RNA sample digested with DNase I can be taken out for conventional PCR reaction. When it was found that no amplified band was generated, the reverse transcription step was performed again, and at the same time, in order to further verify whether the band amplified by real-time quantitative PCR was GmFER, the PCR product was gel-cut and recovered for sequencing. The specific method is as follows: total RNA is extracted from materials such as soybean roots, stems, leaves, f...

Embodiment 3

[0038] Example 3: Construction of yeast expression vector containing GmFER gene and detection of tolerance of transgenic yeast to Cd.

[0039] (1) Construction of yeast expression vector:

[0040] Using the full-length sequence of GmFERcDNA as a template, forward primer P5 (shown in SEQ ID NO: 7, synthesized by Shanghai Bioengineering Co.) and reverse primer P6 (shown in SEQ ID NO: 8) containing restriction sites , synthesized by Shanghai Bioengineering Company), PCR amplified the full-length sequence of GmFERcDNA with BamHI recognition site added at the 5' end and SmaI recognition site added at the 3' end. The nucleotide sequences of the primers were as follows:

[0041] P5: 5′-CGC GGATCC ATGGATGTTCACGAAGACACACTCA-3' (the underlined base is the recognition site of the restriction endonuclease BamHI);

[0042] P6: 5′-TCC CCCGGG TCAAGCAGGAAAAGATGCCACGAAT-3' (bases underlined are recognition sites for restriction endonuclease SmaI).

[0043] After the reaction, the PCR amp...

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Abstract

The invention discloses a soybean bHLH transcription factor gene GmFER and encoded protein and application thereof. The nucleotide sequence of the gene GmFER is as shown in SEQ ID No.1, and the amino acid sequence of the protein encoded by the gene is as shown in SEQ ID No. 2. A semi-quantitative detecting result shows that the gene GmFER is mainly expressed in soybean roots; the expression quantity of the gene GmFER is increased evidently after Cd stress, that is to say, the gene GmFER is induced by Cd; by overexpressing the gene GmFER in yeast and plants, the Cd tolerance of the yeast and plants can be increased, and the gene GmFER is high in application value.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering, and relates to plant FER gene and its coded protein and application, in particular to soybean bHLH transcription factor gene GmFER, its coded protein, and its application in improving the tolerance of yeast and tobacco to cadmium. Background technique [0002] With the increasing development of society, the rapid expansion of non-agricultural construction land, the increase of waste discharge and the use of agricultural chemical fertilizers, soil heavy metal pollution has gradually become a worldwide environmental problem. Cadmium (Cd) is a highly toxic heavy metal with a very long half-life, and excessive intake can lead to kidney and bone lesions (see Nawrot T., Plusquin M., Hogervorst J., et al., Environmental exposure to cadmium and risk of cancer: aprospective population-based study [J], Lancet Oncology, 2006, 7(2):119-126). [0003] Soybean is a crop with high economic value in agr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/70C12N15/81C12N5/10C12N15/11A01H5/00
CPCC07K14/415C12N5/04C12N15/11C12N15/81C12N15/8205C12N2502/70C12N2502/99
Inventor 刘晓庆陈新陈华涛张红梅崔晓艳袁星星
Owner JIANGSU ACAD OF AGRI SCI
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