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Molecular marker in close linkage with cytoplasm male sterile (CMS) restoring gene of capsicum annuum L. and acquisition method and application thereof

A technology for restoring genes and male sterility, which is applied in the fields of pepper genetics and breeding and molecular biology, can solve the problems of pepper breeding lacking cytoplasmic male sterility restoration gene markers, etc., and achieves reduction of field planting scale and later identification workload. The effect of saving manpower and speeding up the breeding process

Active Publication Date: 2018-11-06
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there is still a lack of reliable cytoplasmic male sterility restoration gene markers in pepper breeding. Based on this, this application is made

Method used

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  • Molecular marker in close linkage with cytoplasm male sterile (CMS) restoring gene of capsicum annuum L. and acquisition method and application thereof
  • Molecular marker in close linkage with cytoplasm male sterile (CMS) restoring gene of capsicum annuum L. and acquisition method and application thereof
  • Molecular marker in close linkage with cytoplasm male sterile (CMS) restoring gene of capsicum annuum L. and acquisition method and application thereof

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Experimental program
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Effect test

Embodiment 1

[0045] A molecular marker CaML closely linked with the pepper cytoplasmic male sterility restoration gene of this embodiment, said molecular marker is transformed from the genotyping primer designed at the SNP locus closely linked with the pepper cytoplasmic male sterility restoration gene .

[0046] The molecular marker CaML closely linked to the capsicum cytoplasmic male sterility restorer gene described above is obtained by the following method, including the following steps:

[0047] (1) Group construction:

[0048] The cytoplasmic male sterile line material YJ15(S / rfrf) was used as the female parent, and the restorer line materials XJ02(N / RfRf) and XJ07(N / RfRf) were used as the male parent to construct F2 segregating populations BL and HS, respectively.

[0049] (2) Phenotype identification:

[0050] F2 populations were planted in plastic greenhouses, covered with insect-proof nets, and managed according to the standardized production mode of pepper. By observing wheth...

Embodiment 2

[0058] Example 2 Verification of capsicum cytoplasmic male sterility restoration gene molecular marker CaML:

[0059] Utilize the recovery gene molecular marker primer that embodiment 1 obtains, respectively to the F in embodiment 1 2 Groups BL and HS are identified, and the steps are as follows:

[0060] (1) Using pepper genomic DNA as a template, the concentration is 80-120ng / μl; molecular marker CaML is used for PCR amplification, and the sequence of the forward primer is CaML-F: 5'-TGGCCTTGCAATCCGTTATG-3', and the sequence of the reverse primer is CaML-R: 5'-TGCTAGTGTTACGGGTATGCTC-3'. PCR reaction system: the total volume is 15 μl, and the specific components are as follows: ddH 2 O 10.4μl, 10×PCR Buffer 1.5μl, 1.2μl dNTPs (2mmol / L), 0.08μl Taq DNA polymerase (5U / μl), 0.4μl forward primer (10μM), 0.4μl reverse primer (10μM) and 1.0 μl DNA template.

[0061] (2) PCR amplification program: pre-denaturation at 95.0°C for 3 min; denaturation at 95.0°C for 30 s, renaturatio...

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Abstract

The invention relates to a molecular marker in close linkage with cytoplasm male sterile restoring gene of capsicum annuum L. and an acquisition method and application thereof. A CMS line material YJ15 and a CMS restoring line material XJ07 are utilized to construct a F2 segregating population HS, and meanwhile, the CMS line material YJ15 and a CMS restoring line material XJ02 are utilized to construct the other F2 segregating population BL. Positioning of the restoring gene to the population HS is implemented through an extreme mixer by an RNA-Seq and recombinant monoclonal analysis method toacquire a chromosome region and a candidate gene which are in close linkage with the restoring gene, and further, the molecular marker CaML is designed according to variation near the candidate gene.When the marker is used for identifying the population HS , the coincidence rate reaches 100%; when the marker is used for identifying the population BL, the coincidence rate is up to 93.2%. The molecular marker which can be in close linkage with the CMS restoring gene of capsicum annuum L. is provided for cloning and relative breeding of the CMS restoring gene of capsicum annuum L. and has highvalue in use.

Description

technical field [0001] The invention belongs to the fields of capsicum genetic breeding and molecular biology, and in particular relates to a molecular marker closely linked with a capsicum cytoplasmic male sterility restoration gene, an obtaining method and application thereof. Background technique [0002] Capsicum (Capsicum annuum L.) is an annual or perennial plant belonging to the Solanaceae Capsicum genus, which is widely planted all over the world and is an important vegetable crop. Pepper originated in South America, and its history of artificial domestication and cultivation is more than 6000 years. Around the late Ming Dynasty (16th century), pepper was introduced into my country. It has become an important part of the daily diet of people in my country, especially in the Yangtze River Basin. [0003] How to efficiently breed high-yield and high-quality pepper varieties has always been the concern of pepper breeders. The current pepper breeding in my country is s...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12Q1/6806C12N15/11
CPCC12Q1/6806C12Q1/6895C12Q2600/13
Inventor 欧阳波贾全锐于会洋周玉红刘峰陈文超李峰邹学校
Owner HUAZHONG AGRI UNIV
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