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Abelmoschus esculentus chloroplast microsatellite polymorphic marker primers and method for molecular identification of varieties

A technology of microsatellite polymorphism and molecular identification of varieties, applied in the field of molecular biology, can solve the problems of incomplete identification of okra resources, etc., and achieve the effects of multiple copies, small molecular weight, accurate identification and identification, and simple structure

Active Publication Date: 2019-02-15
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The invention provides a method for identifying germplasm resources of okra using other substances other than nuclear DNA of okra, and solves the incomplete problem in the identification of okra resources

Method used

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  • Abelmoschus esculentus chloroplast microsatellite polymorphic marker primers and method for molecular identification of varieties
  • Abelmoschus esculentus chloroplast microsatellite polymorphic marker primers and method for molecular identification of varieties
  • Abelmoschus esculentus chloroplast microsatellite polymorphic marker primers and method for molecular identification of varieties

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Embodiment 1: okra chloroplast microsatellite molecular marker polymorphic primer (cpSSR) acquisition:

[0063] (1) Chloroplast microsatellite molecular marker sequence screening

[0064] The complete sequence of the chloroplast genome of okra (Abelmoschus esculentus) (NC_035234) was downloaded from the GenBank database (https: / / www.ncbi.nlm.nih.gov / ), the complete sequence of the chloroplast genome of okra was analyzed, and the molecular sequences of chloroplast microsatellites were screened. Use the MISA perl script software (http: / / pgrc.ipk-gatersleben.de / misa / ) to search for SSR sites (simple sequence repeats, SSR) distributed in the okra chloroplast genome, and the parameters are set as follows: single nucleotide repeat sequence (mononucleotide), repeating units greater than or equal to 10; dinucleotide repeating sequences (dinucleotides), repeating units greater than or equal to 5; trinucleotide repeating sequences (trinucleotides), repeating units greater than or...

Embodiment 2

[0094] Embodiment 2: the present embodiment provides a kind of method that utilizes the polymorphic primer of the chloroplast microsatellite molecular marker that embodiment 1 provides to carry out the kinship analysis of okra variety, specifically as follows:

[0095] Sample: 20 parts of sample materials of 5 okra varieties provided by the embodiment of the present invention 1, and extract 20 parts of total genomic DNA of test samples, to detect the polymorphic primers of okra chloroplast microsatellite molecular markers in different varieties of okra versatility.

[0096] Using the polymorphic primers of chloroplast microsatellite molecular markers in okra provided in Example 1 as PCR extension primers, PCR amplification was performed on the total genomic DNA of 20 samples. Separation of the amplified product by non-denaturing polyacrylamide coagulation electrophoresis, if there is a DNA electrophoresis band at the same site, it is recorded as "1", if there is no DNA electro...

Embodiment 3

[0100] Embodiment 3: This embodiment provides a method for molecular identification of okra varieties using the polymorphic primers of chloroplast microsatellite molecular markers provided in embodiment 1. details as follows:

[0101] (1) extract the total genomic DNA of the sample of the tested okra variety;

[0102] (2) With the DNA extracted in step (1) as a template, the polymorphic primers of the okra chloroplast microsatellite molecular markers provided by Example 1 are used as PCR amplification primers to carry out PCR amplification respectively;

[0103] (3) Carrying out 8% non-denaturing polypropylene gel electrophoresis detection to the PCR product of step (2), obtaining the DNA fingerprint of the sample to be tested;

[0104] (4) Carry out band type statistics on the DNA fingerprint of the sample in step (3), and determine the okra species to be tested by comparing and judging specific electrophoretic bands.

[0105] It was found that 4 pairs of cpSSR primers ampl...

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Abstract

The invention discloses abelmoschus esculentus chloroplast microsatellite polymorphic marker primers and a method for molecular identification of varieties. The sequences of the polymorphic marker primers are shown as SEQ ID NO. 1-SEQ ID NO. 24. Based on the provided polymorphic marker primers, the genetic relationship analysis can be performed on the abelmoschus esculentus varieties through conventional PCR (Polymerase Chain Reaction) amplification, non-denatured polyacrylamide gel electrophoresis and DNA fingerprint analysis. Moreover, the abelmoschus esculentus varieties are subjected to PCR amplification and electrophoresis detection by utilizing the provided polymorphic marker primers to obtain four specific DNA sequences of the specific abelmoschus esculentus varieties; based on thespecific DNA sequences, the molecular identification of the target abelmoschus esculentus varieties can be achieved. The polymorphic marker primers and the application method thereof, disclosed by theinvention, can reflect genetic information of cytoplasmic genomes of the abelmoschus esculentus varieties. The invention provides a method for the identification of germplasm resources of abelmoschusesculentus with other substances other than abelmoschus esculentus cell nucleus DNA and solves the problem of incompleteness in the identification of the abelmoschus esculentus resources.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, in particular to okra chloroplast microsatellite polymorphism marker primers and a variety molecular identification method. Background technique [0002] Okra (Abelmoschus esculentus L.) is an annual herb of the genus Okra in the Malvaceae family, also known as okra, ambrette, kidney tonic grass, sheep's horn, sheep sesame, coffee sunflower, etc. It is native to Africa and is now widely cultivated in tropical and subtropical regions. The tender okra fruit is rich in protein, cellulose, unsaturated fatty acids and minerals such as iron, manganese, zinc, calcium and potassium, and has high edible value. , Canning, etc. In addition, it is also rich in polysaccharides, flavonoids, anthocyanins and terpenes and other biologically active ingredients, which have the functions of helping digestion, anti-cancer, anti-tumor, anti-inflammatory, protecting the liver and enhancing human immunity. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/156
Inventor 俞晨良孙晨栋董文其
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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