Masson pine SNP (single nucleotide polymorphism) genotyping method based on HRM (high-resolution melt) technology

A genotyping method and masson pine technology, applied in the field of genetic engineering, to achieve the effects of high sensitivity, reduced use of reagents and consumables, and high comprehensive efficiency

Active Publication Date: 2019-08-16
NANJING FORESTRY UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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  • Masson pine SNP (single nucleotide polymorphism) genotyping method based on HRM (high-resolution melt) technology
  • Masson pine SNP (single nucleotide polymorphism) genotyping method based on HRM (high-resolution melt) technology
  • Masson pine SNP (single nucleotide polymorphism) genotyping method based on HRM (high-resolution melt) technology

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Experimental program
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Embodiment 1

[0077] Embodiment 1 Masson pine HRM-SNP marking method

[0078] 1. Screening of SNP sites of Pinus massoniana

[0079] Based on the transcriptome sequencing results of the differentiated xylem and its protoplasts of Pinus massoniana, the sequences were compared by tools such as samtools and picard-tools, and finally by the variation detection software GATK3 (Van der Auwera GA, Carneiro M O, Hartl C, et al. From FastQ data to high-confidence variant calls: the genome analysis toolkit best practices pipeline [J]. Current protocols in bioinformatics, 2013, 43 (1): 11.10.1-11.10.33.) Mining of SNP Calling sites , and filter the original results (filtering out SNPs with a quality value less than 40 and a distance less than 2), and screen the SNP sites according to the obtained predicted sites and their mutation information. The screening criteria are as follows: the predicted SNP sites are in Different types of base mutations were found in the differentiated xylem and its protopla...

Embodiment 2

[0133] Example 2 Application of the SNP genotyping method of Pine massoniana based on HRM technology in the construction of the SNP fingerprint of Pine massoniana samples

[0134] Based on the genotyping results, the fingerprints of 27 Pine massoniana samples were drawn. As shown in Table 6, the typing results of the same primer were distinguished by different numbers, indicating that each Pine massoniana sample was divided into different groups during genotyping. Since only 6 of them (SNP-2, SNP-3, SNP-4, SNP-5, SNP-6 and SNP-9) can completely distinguish 27 Pinus massoniana and construct DNA fingerprints, fingerprints are not Contains SNP-1, SNP-7, SNP-8 and SNP-10.

[0135] Table 6 Fingerprints drawn by 6 SNP sites on 27 samples of Pine massoniana

[0136] dna sample fingerprint code dna sample fingerprint code dna sample fingerprint code 1 154455 10 125134 19 124134 2 534154 11 424142 20 425144 3 534555 12 425554 21 124542 ...

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Abstract

The invention discloses a masson pine SNP (single nucleotide polymorphism) genotyping method based on the HRM (high-resolution melt) technology, and belongs to the technical field of genetic engineering. The masson pine SNP genotyping method includes screening ten SNP loci, designing corresponding primers, extracting genomic DNA of masson pines, carrying out PCR reaction and HRM analysis, collecting data, performing SNP locus genotype analysis on each sample, and creating SNP fingerprints of the masson pine samples by the method. The masson pine SNP genotyping method has the advantages of simplicity, convenience and rapidness in operation, low cost, accurate result, and capability of being applied to molecular marker assisted breeding and genetic resource diversity research.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a method for SNP genotyping of Pine massoniana based on HRM technology. Background technique [0002] Masson pine is one of the important high-quality timber tree species in my country. It has high economic value and wide application, and occupies an important position in the development of forest resources, industrialization of rosin forest and forest ecological service functions. With the development of scientific and technological means and the advancement of scientific research, the research on Pinus massoniana has gone from the traditional field of morphology and application to the field of molecular biology. [0003] SNP (single nucleotide polymorphism) is the most abundant form of genetic variation in organisms. At present, according to the level of automation, SNP detection methods are roughly divided into two categories, namely, SNP detection meth...

Claims

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Application Information

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IPC IPC(8): C12Q1/6827
CPCC12Q1/6827C12Q2531/113C12Q2565/125C12Q2527/107
Inventor 胥猛祁浩然沈海伦徐梦璇
Owner NANJING FORESTRY UNIV
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