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Method for constructing walnut microsatellite DNA marker fingerprint spectrum and primer pair applied to method

A technology of DNA labeling and fingerprinting, which is applied in the direction of recombinant DNA technology, DNA/RNA fragments, microbial measurement/inspection, etc., can solve the problem of high genetic similarity of varieties, achieve high polymorphism, stable results, and easy and fast retrieval Effect

Pending Publication Date: 2018-07-20
HORTICULTURE INST OF XINJIANG ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

As far as the country is concerned, due to the high genetic similarity of walnut local varieties introduced and utilized between regions, it is difficult for traditional morphological and cytological identification methods to accurately and truly identify the differences in walnut varieties. Analysis and identification

Method used

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  • Method for constructing walnut microsatellite DNA marker fingerprint spectrum and primer pair applied to method
  • Method for constructing walnut microsatellite DNA marker fingerprint spectrum and primer pair applied to method
  • Method for constructing walnut microsatellite DNA marker fingerprint spectrum and primer pair applied to method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Utilize special-purpose primer HSSR2 to construct the method for walnut microsatellite DNA marker fingerprint:

[0021] 1. DNA extraction

[0022] Extract the DNA of biological samples of different walnut varieties, take part of the DNA solution to measure its concentration, dilute to 20ng / μL, and store it at -20°C for later use;

[0023] 2. Use the genomic DNA of walnut in step 1 as a template, and use the special primer HSSR2 to perform PCR amplification; the PCR reaction system is (20μL): 1×Buffer, 1.5mmol / L MgCl 2 , 0.1mmol / L dNTPs, 500nmol / L primer, 1 UTaq enzyme, 50ng DNA template. The reaction was carried out on an Eppendorf Mastercycler gradient PCR amplification instrument. The PCR program was: 94°C for 2min; 94°C for 15s, 52-65°C for 30s, 72°C for 90s, 35 cycles; 72°C for 10min;

[0024] 3. PCR products were separated by electrophoresis on 6.0% polyacrylamide gel and detected.

[0025] Establish a fingerprint map based on the band information presented by t...

Embodiment 2

[0030] Utilize special-purpose primer HSSR4 to construct the method for walnut microsatellite DNA marker fingerprint:

[0031] 1. DNA extraction

[0032] Extract the DNA of biological samples of different walnut varieties, take part of the DNA solution to measure its concentration, dilute to 20ng / μL, and store it at -20°C for later use;

[0033] 2. Use the genomic DNA of walnut in step 1 as a template, and use the special primer HSSR4 to perform PCR amplification; the PCR reaction system is (20μL): 1×Buffer, 1.5mmol / L MgCl 2 , 0.1mmol / L dNTPs, 500nmol / L primer, 1 UTaq enzyme, 50ng DNA template. The reaction was carried out on an Eppendorf Mastercycler gradient PCR amplification instrument. The PCR program was: 94°C for 2min; 94°C for 15s, 52-65°C for 30s, 72°C for 90s, 35 cycles; 72°C for 10min;

[0034] 3. PCR products were separated by electrophoresis on 6.0% polyacrylamide gel and detected.

[0035] Establish a fingerprint map based on the band information presented by t...

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Abstract

The invention relates to a method for constructing a walnut microsatellite DNA marker fingerprint spectrum and a primer pair applied to the method. A special primer HSSR2 for constructing the walnut microsatellite DNA marker fingerprint spectrum is constructed, the sequence of a forward primer is 5'-GATTTCTCCAGGAAGGCTCC-3', and the sequence of a reverse primer is 5'-AGGAAGAAGAGGAGGGCTTG-3'; a specific primer HSSR4 is constructed, the sequence of a forward primer is 5'-GAATGATGGCCTTGATGATTG-3', and the sequence of a reverse primer is 5'-GCAACCCATCCTCAACTTCT-3'. The constructed DNA marker fingerprint spectrum has high polymorphism, abundant fingerprints and good repeatability; the identification accuracy and efficiency of detecting walnut varieties by using the DNA marker fingerprint spectrum are greatly improved.

Description

technical field [0001] The invention relates to a method for constructing a walnut DNA fingerprint and a pair of primers used therefor. Background technique [0002] The hometown of walnut (Juglans regia) is Iran in western Asia. Zhang Qian brought it back to China after his mission to the Western Regions in the Han Dynasty. Therefore, walnuts are also called walnuts and Qiang peaches, which are Juglandaceae plants. Walnuts, almonds, cashews, and hazelnuts are also known as the world-renowned "four major dried fruits". [0003] Walnuts are rich in nutritional value and have the reputation of "longevity fruit", "longevity fruit" and "health treasure". 86% of the fat in walnuts are unsaturated fatty acids, walnuts are rich in copper, magnesium, potassium, vitamin B 6 , folic acid and vitamin B 1 , also contains fiber, phosphorus, niacin, iron, vitamin B 2 and pantothenic acid; every hectogram contains 15-20 grams of protein, more fat, and 10 grams of carbohydrates; Moreo...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/156
Inventor 马凯韩立群李宁梅闯闫鹏艾沙江·买买提王继勋赵国庆张校立
Owner HORTICULTURE INST OF XINJIANG ACAD OF AGRI SCI
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