InDel molecular marker interlocked with gossypium harknessii male sterile recovery gene and application

A Hackney cotton and male sterility technology, applied in the field of genetic engineering, can solve the problems of time-consuming and labor-intensive identification of field materials, poor accuracy, etc., and achieve the effect of eliminating field experiments, high accuracy, and increasing cotton yield

Active Publication Date: 2018-11-02
INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0005] In order to solve the shortcomings of conventional breeding methods, such as time-consuming, labor-intensive and poor accuracy of field material identification in the process of restoration material selection and improvement, to improve the efficiency of identification of materials containing restorer genes, and to ensure the purity of restorer line seeds during the seed production process, the present invention Provides an InDel molecular marker linked to the Hackney cotton male sterility restorer gene and InDel molecular marker primers, which can speed up the process of cotton restorer line selection and improvement, and at the same time ensure the purity of restorer line seeds

Method used

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  • InDel molecular marker interlocked with gossypium harknessii male sterile recovery gene and application
  • InDel molecular marker interlocked with gossypium harknessii male sterile recovery gene and application
  • InDel molecular marker interlocked with gossypium harknessii male sterile recovery gene and application

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Experimental program
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Effect test

Embodiment 1

[0027] We found an InDel sequence in the promoter region of the gene named Gh_D05G3392 on chromosome Gh_D05 of the upland cotton genome. Restorer gene linkage for male sterility traits. In the material containing the restorer gene, there is a 12nt insertion sequence "TAGAAGACTGGA" at this position, and the sequence coding direction is 5'→3' direction; in the material without the restorer gene, this insertion sequence does not exist.

[0028] InDel molecular marker linked to Hackney cotton male sterility restorer gene, such as figure 1 As shown, its sequence is: 5'-TAGAAGACTGGA-3'.

[0029] According to the InDel sequence and PCR primer design principles, primers were designed upstream and downstream, and successfully converted into InDel marker primers. The primer pairs for amplifying the InDel molecular markers are shown in Table 1:

[0030] Table 1: InDel Molecular Labeling Primers

[0031]

[0032] A 206bp band containing the insert sequence was specifically amplified...

Embodiment 2

[0034] 1. Cotton cytoplasmic male sterility restorer gene near-isogenic line material and restorer gene backcross population material (BC 5 f 2 ) Extraction of seed DNA

[0035] Cotton restorer gene near-isogenic line material and restorer gene backcross population material (BC 5 f 2) seeds were shelled, and each seed was crushed separately and then transferred to a 2ml centrifuge tube; 800 μl of DNA lysate (0.05M Tris-Cl, 0.01M EDTA, 2% SDS, 1% PVP, 0.5% sorbitol, 0.705 M NaCl, adjust the pH value to 8.0, autoclave; add 1% β-mercaptoethanol before use), mix well and place in a water bath at 65°C for 30 min, and shake gently every 10 min; after the water bath, add 800 μl of chloroform: isoamyl alcohol (24:1), slowly invert and mix until there is no layering, centrifuge at 12000rpm at 4°C for 10min; use a tipless pipette to draw the supernatant (about 800μl) and transfer it to another 2ml centrifuge tube, add the supernatant Equal volume of chloroform:isoamyl alcohol (24:1)...

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Abstract

The invention discloses an InDel molecular marker interlocked with a gossypium harknessii male sterile recovery gene and application. The molecular marker is 5'-TAGAAGACTGGA-3', and is located at a position, which is distant from an initiation codon by 1247 bp in a promoter region of the recovery series gene Gh_D05G3392, and an InDel molecular marker primer is as shown in SEQID NO. 1-2. By the InDel molecular marker primer, whether recovery genes in materials exist or not and are homozygous or not is effectively identified by operations such as simple PCR and polyacrylamide gel electrophoresis, therefore, breeding of a field cotton cytoplasmic male sterile new recovery series material is accelerated, and seed purity of cotton recovery series is ensured.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and relates to an InDel molecular marker linked with a Hackney cotton male sterility restoration gene and its application. Background technique [0002] Cotton is an important economic crop in my country and has an important position and significance in agricultural production. The research and utilization of heterosis can effectively improve cotton yield and stress resistance. Hybrid seed production is the main manifestation of heterosis utilization at present. The first generation of cotton hybrids shows significant heterosis, which increases yield by about 15% compared with conventional varieties. In actual production, cotton hybrids can be produced by methods such as artificial emasculation, chemical detasseling, and "three-line matching" methods. The artificial detasseling pollination method is time-consuming, labor-intensive and costly, and the purity of the obtained hybrids i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 张兵兵吴建勇邢朝柱郭立平戚廷香张学贤王海林唐会妮乔秀琴
Owner INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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