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Application of sea island cotton gbnac1 in resistance to verticillium wilt

A technology against Verticillium wilt and Verticillium wilt, applied in the field of genetic engineering, can solve problems such as lack of preventive measures

Active Publication Date: 2019-12-03
HENAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Preliminary statistics show that Verticillium wilt is getting worse year by year in cotton producing areas, but there is still a lack of effective preventive measures

Method used

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  • Application of sea island cotton gbnac1 in resistance to verticillium wilt
  • Application of sea island cotton gbnac1 in resistance to verticillium wilt
  • Application of sea island cotton gbnac1 in resistance to verticillium wilt

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] sea ​​island cotton transcription factor GbNAC1 It is the basis of the present invention, so the cloning and obtaining process of the gene will be briefly introduced as follows.

[0045] 1. Electronic cloning

[0046] First, according to the previous transcriptome sequencing in our laboratory, a 1159bp gene fragment (unigene25035_kv-1) annotated as a member of the NAC family was screened, and electronic cloning was performed on the basis of it to obtain a sequence.

[0047] 2. Design primers for PCR amplification

[0048] First, extract RNA and reverse transcribe to obtain cDNA, specifically:

[0049] After the cotton grows cotyledons, extract cotton RNA (use liquid nitrogen grinding method to extract RNA, refer to the instruction of EASYspin Plus Plant RNA Rapid Extraction Kit for specific steps), and reverse transcribe it into cDNA;

[0050] The cDNA template preparation system is specifically designed as follows:

[0051] oligo(dT), 2 μL;

[0052] RNA, 3 μL;

...

Embodiment 2

[0084] for further understanding GbNAC1 The related characteristics of genes, the inventors use specific carrier pairs GbNAC1 Subcellular localization of genes in cells, and the use of GUS gene markers for GbNAC1 The expression of genes in plant tissues was analyzed, and the relevant experimental procedures are briefly described as follows.

[0085] Subcellular localization of GbNAC1 gene in cells

[0086] Using Gateway technology to GbNAC1 The gene was recombined into the subcellular localization vector pK7FWG2.0, and then transformed into Nicotiana benthamiana ( Nicotiana benthamiana ), observed by confocal laser microscopy GbNAC1 The localization and expression of genes in cells. The specific experimental process is:

[0087] Firstly, the primer sequences were designed, and the genomic gDNA extracted from the leaves of cotton Xinhai 16 was used as a template to carry out GbNAC1 Amplification of the sequence, electrophoresis and recovery of the amplified product, the...

Embodiment 3

[0118] This example mainly introduces GbNAC1 The distribution of genes in cotton tissue and the changes of gene expression induced by Verticillium dahliae, the relevant experimental process is briefly described as follows.

[0119] Distribution of GbNAC1 Gene in Cotton Tissues

[0120] for the sake of GbNAC1 The function of the gene has a preliminary understanding, so it is necessary to GbNAC1 The distribution of genes in different tissues has a preliminary understanding, and the relevant experimental processes are mainly:

[0121] After the sea island cotton Xinhai 15 seeds were sown, when two true leaves grew out, the roots, stems, true leaves, cotyledons, and shoot tips were sampled (preserved at -80°C before sample processing), and the RNA was extracted by liquid nitrogen grinding method , and reverse-transcribed into cDNA, which was used as an amplification template for fluorescent quantitative PCR.

[0122] Fluorescent quantitative PCR in different tissue samples Gb...

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Abstract

The invention belongs to the technical field of genetic engineering and particularly relates to application of sea island cotton GbNAC1 gene in verticillium wilt resistance. The GenBank ID of a GbNAC1 gene is KP317496, and the gene is relevant to the verticillium wilt resistance of plants. The inventor finds that when the plants are infected by verticillium dahlia, the expression quantity of the gene is reduced, the verticillium wilt resistance of the plants is reduced after the gene is further silenced, and the verticillium wilt resistance of the plants is improved after the gene overexpression. Based on the characteristics, GbNAC1 overexpression plants can be cultivated or screened, better references can be provided for new varieties of plants having better verticillium wilt resistance, and accordingly the sea island cotton GbNAC1 has a better application value on the aspect of cultivation of new varieties of plants.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, in particular to sea-island cotton GbNAC1 Patent application for the application of genes in Verticillium wilt resistance. Background technique [0002] During the growth and development of plants, they are often subjected to a series of biotic and abiotic stresses such as various pathogenic bacteria or drought, high salinity, and low temperature. These stresses usually cause damage to plants and affect the growth and yield of plants. Transcription factors are a class of important regulatory genes that regulate gene expression. They play a very important role in the process of plants resisting adversity stress. They activate or inhibit the transcription and expression of target genes by binding to specific DNA sequences in the promoters of target genes. There are a large number of genes encoding different types of transcription factors in plants, among which AP2 / ERF, bZIP, WRKY, MYB...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82A01H5/00A01H6/20A01H6/82
CPCC07K14/415C12N15/8282
Inventor 蔡应繁王微娜杨璨耿帅鹏孙全张骁宋纯鹏
Owner HENAN UNIVERSITY
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