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Method for identifying sika deer, red deer or other hybridized deer on basis of COI and SRY sequences

A sika deer, SRY gene technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve problems such as inability to identify hybrid deer

Active Publication Date: 2017-12-29
华润三九现代中药制药有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the COI gene is mitochondrial DNA, which belongs to cytoplasmic inheritance. This molecular identification method based on the mitochondrial sequence of antler can only be used as evidence to identify the female parent, and cannot directly identify hybrid deer.

Method used

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  • Method for identifying sika deer, red deer or other hybridized deer on basis of COI and SRY sequences
  • Method for identifying sika deer, red deer or other hybridized deer on basis of COI and SRY sequences
  • Method for identifying sika deer, red deer or other hybridized deer on basis of COI and SRY sequences

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] Example 1. SNP sites for identifying sika deer, red deer and their hybrid deer and primers for amplifying them

[0078] The SRY gene is a sex-determining gene in mammals, and the location, size, and structure of the SRY gene vary among different species. Therefore, using COI and SRY genes to establish molecular identification of sika deer, red deer and their hybrid deer has become a new research direction.

[0079] 1. Discovery and application of SNP loci for identifying sika deer, red deer and their hybrid deer

[0080] According to the COI and SRY gene sequences of the tested samples obtained by sequencing, the COI and SRY gene sequences of sika deer, red deer, reindeer, sambar and other species were downloaded from the NCBI database, and homologous alignment was performed using ClustulX 2.1 software. Then analyze its specific SNP sites, and screen suitable identification sites from the specific SNP sites.

[0081] After comparison, the 391st position of the COI gen...

Embodiment 2

[0098] Example 2, Establishment of Molecular Identification Method for Antler Samples of Sika Deer, Red Deer and Their Hybrid Deer

[0099] 1. Obtaining the fragment to be detected

[0100] 1) Extraction and purification of genomic DNA

[0101] The velvet antler medicinal sample was taken, and after surface disinfection with 75% ethanol, 20 mg sample powder was drilled with a bone drill, and the decoction pieces were directly crushed. The total DNA of velvet antler was extracted by using TIANDZ column bone DNAout, which was used as a template for the identification of velvet antler DNA.

[0102] 2) Universal primers amplify the COI gene fragment to be detected and the SRY gene fragment to be detected

[0103] Using the total DNA obtained in the above 1) as a template, use the universal primers CO-F and CO-R (see Table 2) to amplify the COI gene sequence. The PCR reaction program is: 95°C, 5min; 95°C, 30s, 45°C, 1.5min, 72°C, 1.5min, 5 cycles; 95°C, 1min, 50°C, 1.5min, 72°C, ...

Embodiment 3

[0125] Embodiment 3, multiplex PCR distinguishes tested sika deer, red deer and its hybrid deer samples

[0126] The tested samples were 8 batches of known sika deer antler samples, 16 batches of known red deer antler samples and 3 batches of horse-flower hybrid antler samples collected in the heavy circulation market.

[0127] The method of Example 2 was used to carry out multiplex PCR detection on the test samples.

[0128] If the multiplex PCR system for amplifying COI amplifies a specific fragment of 232bp, and the multiplex PCR system for amplifying SRY amplifies a specific fragment of 803bp, then the velvet sample to be tested comes from or the candidate is from sika deer; otherwise, the velvet to be tested The sample does not originate or the candidate does not originate from sika deer;

[0129] If the multiplex PCR system for amplifying COI amplifies a specific fragment of 518bp, and the multiplex PCR system for amplifying SRY amplifies a specific fragment of 425bp, t...

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Abstract

The invention discloses a method for identifying sika deer, red deer or other hybridized deer on basis of COI and SRY sequences. The invention provides the application of a substance for detecting a target site in gene sequences corresponding to a COI gene and an SRY gene in a genome of a to-be-detected sample as well as target nucleotide of the site to preparation of products for detecting whether the to-be-detected sample is sika deer, red deer or other hybridized deer. Two multiple PCR systems are established by utilizing the SRY gene of Y chromosome as an identification mark of a male parent and combining a female parent mark gene COI on the basis of the COI and SRY gene sequences and are used for identifying male and female parent sources of cartialgenous samples, the identification method of the hybridized cartialgenous is initially established, and male and female parent identification of the hybridized deer can be conducted effectively. The method has actual application value on identification of the hybridized cartialgenous, and provides an effective identification method for subsequent quality evaluation of the hybridized cartialgenous.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for establishing identification of sika deer, red deer or hybrid deer based on COI and SRY sequences. Background technique [0002] Antler is one of the traditional famous medicines in my country, which was first recorded in "Shen Nong's Herbal Classic". It has the functions of promoting growth and development, strengthening the body, anti-inflammatory, immune regulation, anti-aging, and promoting wound healing. "Chinese Pharmacopoeia" stipulates that the base of velvet antler is the unossified and densely hairy young horns of the male deer Cervus nippon Temminck or the red deer C. elaphus Linnaeus. [0003] Hybridization is beneficial to increase livestock production. The use of heterosis between species and subspecies of antler deer in my country has always been at the international leading level. As early as 1958, the Longtan Mountain Deer Farm in Jilin City, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/686C12Q1/6888C12Q2600/156C12Q2600/16C12Q2537/143
Inventor 黄璐琦袁媛蒋超魏艺聪赵玉洋
Owner 华润三九现代中药制药有限公司
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