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Indian truffle ssr primer and its cross-species screening method and application

An Indian truffle and product technology, applied in the field of molecular biology, can solve problems such as unseen population genetics research reports and difficulty in group sampling, and achieve the effects of shortening the analysis cycle, being convenient to use, and improving research efficiency

Inactive Publication Date: 2015-11-04
KUNMING INST OF BOTANY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The Indian truffle complex studied by the applicant is mainly distributed in the southwestern region of my country. Due to the difficulty in sampling this group, there are no reports on the population genetics of this group at home and abroad.

Method used

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  • Indian truffle ssr primer and its cross-species screening method and application
  • Indian truffle ssr primer and its cross-species screening method and application
  • Indian truffle ssr primer and its cross-species screening method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Screening of SSR primers for Indian truffle (Tuber indicum):

[0060] 1. SSR primer design

[0061] ①Download the whole genome sequence of Tuber melanosporum from NCBI (http: / / www.ncbi.nlm.nih.gov) database.

[0062] ② Use SSRHunter1.3 to search for simple repeat sequence information sites in the genome sequence.

[0063] ③ Select the sequence with 2 to 6 nucleotide repeat motifs at the information site as the target sequence, use the software Primer Premier5.0 to design SSR primers, and use Oligo6.65 to verify the parameters of the primers. Each parameter meets the following conditions: GC The content is 45%-60%; the annealing temperature is 50-60°C; the expected fragment length is between 100-300bp; the primer length is between 18-25bp.

[0064] ④ Select primer sequences that meet certain conditions for synthesis.

[0065] 2. Primer amplifiability detection

[0066] By repeatedly exploring the PCR reaction system and amplification conditions, the following steps w...

Embodiment 2

[0077] Genetic diversity analysis of Indian truffles based on SSR markers:

[0078] Using the SSR polymorphism primers screened in Example 1 to carry out genetic diversity analysis of Indian truffles, the steps are as follows:

[0079] The Indian truffle samples used in the examples were all collected from Yunnan and Sichuan, China.

[0080] 1. Analyze the source of the sample

[0081] The samples used for the analysis were collected from wild Indian truffle populations in Yunnan and Sichuan. The specific geographic information is as follows: Qujing City, Yunnan (103°17′28.05″E, 26°24′46.84″N); Chuxiong Prefecture, Yunnan (101° 9′5.14″E, 25°35′54.49″N); Baoshan City, Yunnan Province (99°19′41.64″E, 25°12′18.34″N); Nujiang Prefecture, Yunnan Province (98°37′57.50″E, 28° 1′17.41″N); Panzhihua, Sichuan (101°37′58.18″E, 26°22′48.77″N).

[0082] 2. DNA extraction of Indian truffle samples

[0083] Use the improved 4×CTAB method to extract the total DNA of the sample, the specif...

Embodiment 3

[0108] Using SSR primers to construct the genetic map of Indian truffle, locate functional genes and establish germplasm resource bank.

[0109] Individuals of the same species of truffles with different geographical distributions usually show certain differences in phenotypic characteristics and taste attributes. Whether these differences are affected by environmental factors (symbiotic tree species, soil, etc.) or determined by genes is still controversial. SSR markers can analyze a large number of individuals with different geographical origins, thereby constructing a genetic map of Indian truffles, and locating genes that affect truffle ascocarp fruit shape and aroma, while certain genotypes are only in populations in specific regions If it exists, germplasm resource identification and preservation can be carried out for materials from different sources, and a corresponding germplasm resource bank can be established. For example, the 170bp gene in SSR24 only exists in the ...

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Abstract

The invention provides a tuber indicum SSR (Simple Sequence Repeat) primer as well as a trans-species screening method and application thereof. The primer has the advantages of uniform distribution, stable amplification, clear electrophoretic band and rich polymorphism. According to the invention, genetic diversity analysis, germplasm resource appraisal, construction of a genetic map and research of a functional gene can be simply and quickly carried out on the tuber indicum. Compared with the conventional method, an analysis period can be greatly shorted, and efficiency can be improved.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to an Indian truffle (Tuber indicum) SSR primer and a cross-species screening method and application thereof. Background technique [0002] Microsatellite markers (microsatellite), also known as simple sequence repeats (simple sequence repeats, SSR) are widely used in population genetics, hybrid speciation and parental analysis and other research fields. Highly polymorphic microsatellites can also be used for individual identification. Microsatellite loci are usually amplified by PCR, and the amplified products are detected by electrophoretic analysis (eg, fluorescent labeling, silver staining) and separation of alleles by size. Microsatellite DNA is neutral, microsatellite DNA markers are co-dominant Mendelian inheritance, can identify heterozygotes, and are very beneficial to the selection of recessive traits, and SSR markers have no epistasis effect and are...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12Q1/04C12N15/11C12R1/645
CPCC12Q1/686C12Q2531/113C12Q2565/125
Inventor 乔鹏刘培贵田伟陈娟万山平赵文青
Owner KUNMING INST OF BOTANY - CHINESE ACAD OF SCI
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