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Kit and method for identifying Gansu zokor (E.cansus)

A kit and reagent technology, applied in the field of kits for identifying Gansu zokor, can solve problems such as difficulty in accurate identification of species, and achieve the effects of low sequencing length requirements and accurate and reliable results.

Pending Publication Date: 2021-09-28
CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the similar living environment and similar morphology of different zokors, it is very difficult to accurately identify species through morphology without professional training.

Method used

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  • Kit and method for identifying Gansu zokor (E.cansus)
  • Kit and method for identifying Gansu zokor (E.cansus)
  • Kit and method for identifying Gansu zokor (E.cansus)

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Experimental program
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Effect test

Embodiment 1

[0044] Embodiment 1, the present invention distinguishes the kit of Gansu zokor

[0045] The components of the kit of the present invention include:

[0046] (1) PCR amplification reagents: comprising primer pairs of SEQ NO: 1-2 and primer pairs of SEQ NO: 3-4; (2) Reagents for sequencing.

Embodiment 2

[0047] Embodiment 2, PCR primer design and verification of SNP site

[0048] The applicant compared the mitochondrial genome sequences of 121 individual zokors from 8 zokor species, and found that there were 2 Gansu zokor species-specific SNP genotypes in the 12S rRNA gene, and 2 Gansu zokor species-specific SNPs in the 16S rRNA gene genotype. After comparing the two ends of the four SNP sites, it was found that there are conserved sequences, which can be used to design PCR primers, so as to determine the 12S rRNA and 16S rRNA gene fragments as DNA barcodes for Gansu zokor species identification.

[0049] 121 individual zokors from 8 zokor species: 12 prairie zokors, 10 northeast zokors, 15 Chinese zokors, 18 Schneider zokors, 16 Roche zokors, 14 plateau zokors, Gansu zokors There were 24 rats and 12 Qinling zokors.

[0050] The 12S rRNA, 16S rRNA and nearby gene sequences were designed to contain species-specific SNP genotype primers in the conserved region as follows:

[...

Embodiment 3

[0061] Embodiment 3, Zokor species identification in Gansu

[0062] First, the above-mentioned 12S rRNA gene fragment primer pair (SEQ ID NO:1 and SEQ ID NO:2) and 16S rRNA gene fragment primer pair (SEQ ID NO:3 and SEQ ID NO:4) were synthesized.

[0063] Use the following methods to identify:

[0064] a) Using the Qiagen DNeasy Blood&Tissue Kit kit, extract the total genomic DNA of zokor muscle tissue numbered FFX-5, the total genomic DNA of zokor liver tissue numbered ZX-6, and the zokor numbered LD-3 Genomic total DNA from mouse liver tissue;

[0065] b) Using the zokor genome total DNA numbered FFX-5 and ZX-6 described in step a) as a template, use the 12SrRNA and 16S rRNA primer pairs to perform PCR reactions respectively, FFX-5 reaction system 25 μL, annealing temperature 56°C; ZX-6 reaction system 25μL, annealing temperature 55°C; use the total genomic DNA numbered LD-3 as described in step a) as a template, use the 12SrRNA primer pair to perform PCR reaction, reactio...

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Abstract

The invention provides a kit and a method for identifying Gansu zokor ((E.cansus), and belongs to the technical field of molecular identification. The kit comprises a reagent for detecting any one or more of the following four SNP sites in a 12S rRNA gene and a 16S rRNA gene of a species to be detected: the 4th site of a conserved motif sequence as shown in SEQ ID NO: 5, the 25th site of the conserved motif sequence as shown in SEQ ID NO: 6, and the 22th site and the 31st site of the conserved motif sequence as shown in SEQ ID NO: 7. The invention provides the kit for simply, conveniently and accurately identifying the species of the Gansu zokor and the method for identifying the Gansu zokor, the problem of identifying the species of the zokor through morphology is solved, and the kit has an excellent application prospect in species identification of the zokor.

Description

technical field [0001] The invention belongs to the technical field of molecular identification, and in particular relates to a kit and a method for identifying Gansu zokors. Background technique [0002] Zokor is a general term for animals in the subfamily Myospalacinae, mainly distributed in China and southern Siberia. Existing zokors include the genus Eospalax and the genus Myspalax, and almost all extant zokor species are distributed in my country. Flat-headed zokors include 2-3 species such as prairie zokor (M.aspalax) and northeast zokor (M.psilurus). There are 6 species of zokor (E.smithi), zokor (E.rothschildi), plateau zokor (E.baileyi), Gansu zokor (E.cansus) and Qinling zokor (E.rufescens). Zokors live in closed underground tunnel systems almost all their lives, and their unique underground lifestyle makes them special in foraging, mating, breeding and other aspects, which has aroused widespread interest of scientists. [0003] Gansu zokors are distributed in t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/6869
CPCC12Q1/6888C12Q1/6869C12Q2600/156C12Q2600/158C12Q2600/178C12Q2531/113C12Q2535/101C12Q2565/125Y02A40/81
Inventor 张同作蔡振媛邓艳芳张婧捷李全亮高红梅江峰徐波李斌
Owner CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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