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Method for constructing Prunus mume Sieb.et Zucc SSR (simple sequence repeat) genetic map

A technology of genetic maps and construction methods, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc. Marking and other issues to achieve the effect of speeding up the process

Active Publication Date: 2012-10-03
BEIJING FORESTRY UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the lack of a large number of polymorphic molecular markers, the research on its genetics and genomics lags behind other plants in the Rosaceae
[0003] At present, in the genetic analysis of woody ornamental flowers, molecular markers such as RAPD, ISSR, and AFLP are widely used. However, most of these markers are developed by technologies without full genome sequence information. Although they have certain application value, But strong randomness and poor stability

Method used

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  • Method for constructing Prunus mume Sieb.et Zucc SSR (simple sequence repeat) genetic map
  • Method for constructing Prunus mume Sieb.et Zucc SSR (simple sequence repeat) genetic map
  • Method for constructing Prunus mume Sieb.et Zucc SSR (simple sequence repeat) genetic map

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] The construction of embodiment 1 plum blossom genetic mapping population

[0028] According to the principle of parent selection for mapping, the plum blossom variety 'Fenpetal' with a high seed setting rate and well-developed stigma was used as the female parent, and the plum blossom variety 'Button Yudie' with a large amount of loose pollen and high pollen viability was used as the male parent.

[0029] During the early flowering stage and the full flowering stage of the male parent, choose the flower buds of the big bud and the middle bud stage on the branches without damage by diseases and insect pests and physiological defects, peel off the anthers and spread them on sulfuric acid paper, room temperature (25 ℃) humidity (below 35%), Naturally loose powder for 20 hours and store at 4°C for later use. At the initial flowering stage of the female parent, select the short and medium flowering branches that grow robustly, trim them appropriately, and carefully remove th...

Embodiment 2

[0030] Example 2 Acquisition of the SSR core primer set developed based on the whole genome sequence of Meihua

[0031] 1.1 Development of 670 pairs of SSR primers based on the whole genome sequence of Meihua

[0032] On the basis of using the Solexa sequencing technology of Illiumina to complete the whole genome sequence of plum blossom, use the computer program MISA (MIcroSAtellites identification tool, http: / / pgrc.ipk-gatersleben.de / misa) to search for 1- For microsatellite markers of 6 nucleotide repeat units, the standards adopted are as follows: the minimum repeat length of a single nucleotide repeat unit is 10 bp, the minimum repeat length of a dinucleotide to tetranucleotide repeat unit is 12 bp, and the minimum repeat length of a pentanucleotide repeat unit is 12 bp. The minimum repeat length of the nucleotide repeat unit is 15 bp, and the minimum repeat length of the hexanucleotide repeat unit is 18 bp. In short, there are 184629 microsatellite markers in the whole g...

Embodiment 3

[0043] Example 3 Construction of Meihua SSR Genetic Mapping

[0044] 1.1 Extraction of parental genomic DNA

[0045] From the F1 population obtained in Example 1, 190 trees were randomly selected to construct the SSR genetic map. Take 1-3 young leaves below the parietal leaves of the parents and their progeny, and extract the total DNA of the leaves with the Plant Genome Extraction Kit (DP305-02) of Tiangen Company, so that the DNA concentration reaches 50 ng / μl.

[0046] 1.2 PCR amplification using 144 pairs of SSR core primer sets

[0047] (1) Fluorescently label the upstream primers of the screened 144 pairs of SSR core primer sets with FAM, HEX and TEMRA;

[0048] (2) Using fluorescently labeled SSR primers, the parents and their 190 offspring randomly selected for the construction of genetic maps were amplified by PCR;

[0049] Reaction system (10 μL): 10× buffer (containing Mg 2+ ) 1μL, 2.5mM dNTP 0.8μL, Taq DNA polymerase (Promega, Madison, WI, USA) 1U, 10mmol L -1...

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Abstract

The invention provides a method for constructing a Prunus mume Sieb.et Zucc SSR (simple sequence repeat) genetic map, wherein the method is based on a primer sequence of a SSR primer group developed on Prunus mume Sieb.et Zucc whole-gene group sequence information and an amplification method, a primer group comprises 670 pairs of SSR primers; 144 pairs of core primers are selected from the 670 pairs and are used for constructing the Prunus mume Sieb.et Zucc genetic map. According to the invention, a Prunus mume Sieb.et Zucc SSR molecule marking technology system is established; an obtained SSR core primers group can reflect stability and codominance and has advantages of generally existing in gene groups, convenient calculation and the like. The core primers group provided by the invention can be used for research for constructing the Prunus mume Sieb.et Zucc genetic map, analyzing genetic diversity, identifying variety, processing identification, and carrying out molecular marker-assisted breeding and the like; therefore, development of Prunus mume Sieb.et Zucc genetics and genomics is promoted.

Description

technical field [0001] The invention relates to the fields of molecular biology, genetics and genomics, in particular to a method for constructing a plum blossom SSR genetic map. Background technique [0002] Plum blossom (Prunus mume Sieb.et Zucc.) belongs to the genus Prunus in the family Rosaceae. It is native to Sichuan, Yunnan, Tibet, Guizhou and other regions in my country. It has been cultivated for more than 3,000 years and is an important woody ornamental flower in my country. However, due to the lack of a large number of polymorphic molecular markers, the research on its genetics and genomics lags behind other plants in the Rosaceae. [0003] At present, in the genetic analysis of woody ornamental flowers, molecular markers such as RAPD, ISSR, and AFLP are widely used. However, most of these markers are developed by technologies without full genome sequence information. Although they have certain application value, But strong randomness and poor stability. The si...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 张启翔孙丽丹程堂仁杨炜茹王佳禄久幸李旭东
Owner BEIJING FORESTRY UNIVERSITY
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