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Specific primer system of EST (expressed sequence tag)-SSR (simple sequence repeat) molecular markers for Pleurotus ostreatus and application of specific primer system

A technology of specific primers and molecular markers, applied in the field of molecular biology, can solve the problem of lagging research on the genomics of edible and medicinal fungi, and achieve simple and fast genetic diversity analysis, improve research efficiency, and shorten the analysis period.

Inactive Publication Date: 2011-09-14
INST OF AGRI RESOURCES & ENVIRONMENT SHANDONG ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] However, due to the relative lag in the genomics research of edible and medicinal fungi, EST-SSR marker technology is rarely studied in edible and medicinal fungi, and there are only research reports on Lentinus edodes and Pleurotus shimeji, but it has not yet been carried out in Pleurotus spp. research in this area

Method used

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  • Specific primer system of EST (expressed sequence tag)-SSR (simple sequence repeat) molecular markers for Pleurotus ostreatus and application of specific primer system
  • Specific primer system of EST (expressed sequence tag)-SSR (simple sequence repeat) molecular markers for Pleurotus ostreatus and application of specific primer system
  • Specific primer system of EST (expressed sequence tag)-SSR (simple sequence repeat) molecular markers for Pleurotus ostreatus and application of specific primer system

Examples

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Effect test

Embodiment 1

[0048] Example 1: Screening of EST-SSR primers.

[0049] 1. Source and screening of SSR loci

[0050] From the NCBI (http: / / www.ncbi.nlm.nih.gov) database, search and download (in FASTA format) the existing EST sequence of Pleurotus pylori, and use EST-trimmer software to remove polyA or polyA at the 5' or 3' end polyT, remove EST sequences with a length less than 100bp, and retain 700bp at the 5' end of ESTs with a length greater than 700bp; then use CAP3 software for fragment contig analysis and clustering, and the initial assembly parameters set during splicing are all default values, To obtain high-quality Unigenes, use the SSR Finder software to search for SSRs in them. The search criteria are: the minimum repeats of single, di, tri, tetra, penta, and hexanucleotides are 12, 6, 5, 5, 5, and 5, respectively. times, and also includes incompletely repeated SSRs interrupted by a sequence in the middle (not exceeding 10bp).

[0051] 2. Design of EST-SSR labeled primers

[0...

Embodiment 2

[0064] The application of the EST-SSR molecular marker-specific primer system described in Example 1 in the analysis of Pleurotus pachyrhiza genetic diversity and the identification of germplasm resources, the steps are as follows:

[0065] 1. Extraction of DNA from Pleurotus pachyrhiza

[0066] Select Pleurotus 002 strain, Pleurotus 091 strain, Pleurotus 095 strain, Pleurotus 149 strain and Pleurotus 278 purchased from the Institute of Agricultural Resources and Agricultural Regional Planning, Chinese Academy of Agricultural Sciences. No. strain, Pleurotus No. A184 strain, No. A228 strain of Pleurotus, No. 004 strain of Pleurotus, No. 265 strain of Pleurotus, No. 279 strain of Pleurotus, No. A247 strain of Pleurotus Species, Pleurotus 208, Pleurotus 230, Pleurotus A289, Pleurotus A240, Pleurotus A242, using PDA solid medium, at 25 Cultivate at constant temperature for 8 days. The steps of extracting DNA are: (1) Scrape 0.2g of mycelium and grind it with liquid nitrogen, add...

Embodiment 3

[0078] Application as described in Example 2, the difference is that the PCR amplification step has increased the amplification results of the specific primer GL26 on 16 Pleurotus pylori strains, the results are as follows Figure 6 shown. After genetic diversity analysis and polymorphism statistical analysis, the results are as follows: Figure 8 shown.

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Abstract

The invention relates to a specific primer system of EST (expressed sequence tag)-SSR (simple sequence repeat) molecular markers for Pleurotus ostreatus and application of the specific primer system, belonging to the technical field of molecular biology. The specific primer system of EST-SSR molecular markers for Pleurotus ostreatus comprises 4 pairs of specific primers for EST-SSR molecular markers. The invention also provides the application of the specific primer system of EST-SSR molecular markers in the genetic diversity analysis and the germplasm resource identification of the Pleurotusostreatus. The specific primer system disclosed by the invention is used for establishing a core germplasm bank of the Pleurotus ostreatus, has convenience for use, can simply and rapidly carry out the genetic diversity analysis of germplasm resources, germplasm resource identification, genetic map construction and research on functional genes; and compared with a traditional method, the invention can greatly shorten an analytical cycle and increase research efficiency.

Description

technical field [0001] The invention relates to an EST-SSR molecular marker-specific primer system of Pleurotus patellata and its application, and belongs to the technical field of molecular biology. technical background [0002] Pleurotus officinalis is the largest cultivated species of edible fungi in my country. In people's long-term production practice, abundant and precious Pleurotus germplasm resources have been accumulated, especially in recent years, with the rapid development of the Pleurotus cultivation industry, the germplasm resources of Pleurotus is also expanding. According to incomplete statistics, there are currently more than 300 cultivars of Pleurotus pachyrhiza in my country (including strains imported from abroad). Abundant germplasm resources have laid the foundation for the improvement of Pleurotus pachyrhiza, selection of new varieties and genetic engineering. However, the germplasm resource bank of Pleurotus pachyrhiza continues to expand, and it is...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11
Inventor 姚强宫志远高兴喜张雪梅韩建东任鹏飞万鲁长任海霞赵军胜李瑾曲玲
Owner INST OF AGRI RESOURCES & ENVIRONMENT SHANDONG ACADEMY OF AGRI SCI
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