Search

CN-121992113-A - SNP (Single nucleotide polymorphism) marker primer pair, kit, application and method for distinguishing Mandarin fish populations in Heilongjiang and Beijing water systems

CN121992113ACN 121992113 ACN121992113 ACN 121992113ACN-121992113-A

Abstract

The invention discloses a SNP (single nucleotide polymorphism) marker primer pair, a kit, application and a method for distinguishing a Mandarin fish population in Heilongjiang and Beijing water system, which are disclosed by the invention, wherein a locus is lgr p.D874V, is positioned at 14537718 base of chromosome 1, has a base mutation type of T/A, shows different genotypes in the two populations, has obvious population genetic differentiation characteristics through population verification, and can be used as an effective molecular marker for subsequent population identification. Based on the method, the invention also discloses the SNP marker primer pair, a kit containing the primer pair and application of the SNP marker in molecular tracing of mandarin fish groups, germplasm resource identification or group genetic characteristic analysis, and also discloses a method for distinguishing the sources of the mandarin fish groups in Heilongjiang and Beijing water systems and a molecular monitoring method of germplasm resources in mandarin fish places.

Inventors

  • CUI RAN
  • ZHANG XIN
  • HU HONGXIA
  • YU HUANHUAN
  • SONG HAILIANG
  • SUN YANSHENG

Assignees

  • 北京市农林科学院

Dates

Publication Date
20260508
Application Date
20260227

Claims (10)

  1. 1. A SNP marker primer pair for distinguishing a black longjiang and Beijing water system mandarin fish population is characterized in that the locus of the SNP marker is lgr p.D874V, which is positioned at 14537718 base of chromosome 1, and the base mutation type is T/A; the primer pair is a primer pair capable of specifically amplifying a nucleic acid fragment containing the site of the SNP marker.
  2. 2. The SNP marker primer pair for distinguishing between the groups of Mandarin fish in Heilongjiang and Beijing water system according to claim 1, wherein the nucleotide sequence of the primer pair is as follows: the forward primer is 5'-CTCTCCTCTCTGTCTGCCCT-3', and the forward primer is a primer, The reverse primer was 5'-CTGAACCTGTTCCGAGCTGT-3'.
  3. 3. A kit for distinguishing a population of siniperca chuatsi in the black longjiang and beijing river, comprising the SNP marker primer pair for distinguishing a population of mandarin fish in the black longjiang and beijing river according to claim 1 or 2.
  4. 4. The kit for distinguishing mandarin fish populations of claims 3, further comprising one or more of DNA extraction reagents, PCR reaction solutions, and reagents required for sequencing.
  5. 5. The application of the SNP marker of the mandarin fish is characterized in that the locus of the SNP marker is lgr p.D874V, which is positioned at 14537718 base of chromosome 1, and the base mutation type is T/A; the application comprises molecular tracing of mandarin fish groups, germplasm resource identification or group genetic characteristic analysis; the mandarin fish group comprises a black dragon river system mandarin fish group and a Beijing river system mandarin fish group.
  6. 6. A method for distinguishing the population sources of the siniperca chuatsi in Heilongjiang and Beijing water systems is characterized by comprising the following steps of (1) providing a siniperca chuatsi sample to be detected, (2) detecting the genotype of a SNP mark locus in the sample, wherein the SNP mark locus is lgr p.D874V, is positioned at a 14537718 base of chromosome 1, and has a T/A base mutation type, and (3) judging the population source of the siniperca chuatsi to be detected according to the detected genotype.
  7. 7. The method for distinguishing sources of siniperca chuatsi in black longjiang and beijing water system according to claim 6, wherein the locus of the SNP marker is TT type in dominant genotype of the siniperca chuatsi in beijing water system, the genotype frequency is 100%, and TT/TA/AA genotype is expressed in the siniperca chuatsi in black longjiang water system.
  8. 8. The method for distinguishing between the sources of the black-and-white water-based mandarin fish populations according to claim 6 or 7, wherein the SNP marker primer pair of claim 1 or 2 or the kit of claim 3 or 4 is used in the detection step of step (2); And/or, in the detecting step of step (2), determining the genotype of the site of the SNP marker by sequencing the amplified product; and/or, in the step (3), judging the population sources of the mandarin fish to be detected according to the detected genotype comprises the following steps: If the genotype is TT, the individual is indicated to be possibly derived from the Beijing water system mandarin fish group or the genetic background is close to the Beijing water system mandarin fish group, and if the genotype is TA or AA, the individual is indicated to be possibly derived from the Heilongjiang water system mandarin fish group or have genetic penetration of the Heilongjiang water system mandarin fish group.
  9. 9. A molecular monitoring method for regional germplasm resources of mandarin fish is characterized by comprising the following steps: (1) Providing a mandarin fish sample to be tested; (2) Detecting the genotype of a locus of an SNP marker in the sample, wherein the locus of the SNP marker is lgr p.D874V, is positioned at 14537718 base of chromosome 1, and has a base mutation type of T/A; the mandarin fish population comprises a black dragon river system mandarin fish population and a Beijing river system mandarin fish population.
  10. 10. The method for molecular monitoring of germplasm resources in mandarin fish according to claim 9, characterized in that the SNP marker primer pair of claim 1 or 2 or the kit of claim 3 or 4 is used in the detection step of step (2), and/or the genotype of the site of the SNP marker is determined by sequencing the amplified product in the detection step of step (2).

Description

SNP (Single nucleotide polymorphism) marker primer pair, kit, application and method for distinguishing Mandarin fish populations in Heilongjiang and Beijing water systems Technical Field The invention relates to the technical field of aquatic animal molecular genetics and germplasm resource identification, in particular to a SNP (single nucleotide polymorphism) marker primer pair, a kit, application and a method for distinguishing a siniperca chuatsi population in Heilongjiang and Beijing water systems. Background The excellent mandarin fish germplasm resource is the fundamental guarantee of the high-quality development of modern characteristic freshwater fishery, and has key significance for improving the industrial benefit and promoting stable yield and income. Therefore, the system improves the meat quality of the native mandarin fish in China, and becomes one of the key directions of the current mandarin fish genetic breeding research. The siniperca chuatsi in the Heilongjiang river system has excellent quality due to the unique natural geographic environment, and is honored as one of four Chinese fresh water fish. Traditional population discrimination methods rely on morphological features and geographical distribution records, but these methods are susceptible to environmental impact, are subjective, and are difficult to identify individuals of hybrid or confounding origin. The molecular marking technology provides a stable and objective means for group identification. Among them, single Nucleotide Polymorphism (SNP) markers have become powerful tools for population genetics and germplasm identification research because of their advantages of abundant numbers, wide distribution, high stability, easy realization of high-throughput detection, etc. At present, specific SNP markers for effectively distinguishing different geographical groups of mandarin fish have not been reported yet. The development of the markers is helpful for establishing a rapid and accurate molecular identification system, and is used for the healthy development and the seed industry vibration of the mandarin fish industry. Disclosure of Invention The invention aims to provide a SNP marker primer pair, a kit, application and a method for distinguishing the siniperca chuatsi population of the Heilongjiang river and the Beijing water system, so that the siniperca chuatsi population of the Heilongjiang river and the Beijing water system can be effectively distinguished. In order to solve the technical problems, the following technical scheme is adopted: On one hand, the invention provides an SNP marker primer pair for distinguishing a Mandarin fish population in Heilongjiang and Beijing water system, wherein the SNP marker locus is lgr p.D874V, is positioned in a Mandarin fish lgr gene, the sequence of the lgr gene is shown as SEQ ID NO. 1 and is positioned at 14505615-14540530 of a first chromosome, the SNP marker locus is positioned at 14537718 bases of chromosome 1, the base mutation type is T/A, and the locus belongs to a T or A nucleotide sequence polymorphism locus, namely, the corresponding homozygous genotype has two types of TT and AA genotypes. The primer pair is a primer pair capable of specifically amplifying a nucleic acid fragment containing the site of the SNP marker. The SNP marker locus is located in the coding region of a gene lgr of mandarin fish (G protein coupled receptor 4 gene rich in leucine repetitive sequence), the sequence of lgr gene is shown as SEQ ID NO. 1, and the SNP marker locus is located at 14505615-14540530 of a first chromosome. This mutation results in the encoded amino acid at position 874 being changed from aspartic acid (Asp, D) to valine (Val, V) (designated p.D874V). Further, the nucleotide sequences of the primer pairs are: the forward primer (SEQ ID NO: 2:lgr4-F1) is 5'-CTCTCCTCTCTGTCTGCCCT-3'; The reverse primer (SEQ ID NO: 3:lgr4-R1) is 5'-CTGAACCTGTTCCGAGCTGT-3'. On the other hand, the invention also provides a kit for distinguishing the Mandarin fish groups in the Heilongjiang and Beijing water systems, which comprises the SNP marker primer pair for distinguishing the Mandarin fish groups in the Heilongjiang and the Beijing water systems. Further, one or more of DNA extraction reagent, PCR reaction liquid and reagent required for sequencing are also included. In still another aspect, the invention also provides application of the SNP marker of the mandarin fish, wherein the locus of the SNP marker is lgr p.D874V, which is positioned at 14537718 base of chromosome 1, and the base mutation type is T/A; the application comprises molecular tracing of mandarin fish groups, germplasm resource identification or group genetic characteristic analysis; the mandarin fish group comprises a black dragon river system mandarin fish group and a Beijing river system mandarin fish group. On the other hand, the invention also provides a method for distinguishing the population sources of the siniperca ch