Method for identifying parent-child relationship of tetraploid crassostrea gigas based on SNP (Single Nucleotide Polymorphism) sites

Abstract

The invention discloses a tetraploid crassostrea gigas parent-child relationship identification method based on SNP sites, and belongs to the technical field of tetraploid crassostrea gigas molecular assisted breeding. The method for identifying the parent-child relationship of tetraploid crassostrea gigas comprises the following steps: (1) sequencing adductor muscles of tetraploid crassostrea gigas to be detected; (2) comparing the reference genome of the crassostrea gigas to obtain the genotype of the specified SNP site of the tetraploid crassostrea gigas; (3) calculating IBS and LOD of the to-be-detected parents and the to-be-detected offspring according to an SNP typing result; and (4) identifying the parent-child relationship between the to-be-detected parent and the filial generation by combining IBS and LOD. The SNP loci have the advantages that 1000 SNP loci for identifying the parent-child relationship of the tetraploid crassostrea gigas are disclosed for the first time, data support is provided for identifying the parent-child relationship of the tetraploid crassostrea gigas, a foundation is laid for developing a liquid phase chip for identifying the parent-child relationship of the tetraploid crassostrea gigas, and the SNP loci have good application prospects.

Description

Technical Field

[0001] The present invention relates to a parent-offspring identification method, in particular to a parent-offspring identification method of a tetraploid long oyster based on a single nucleotide polymorphism (SNP) site, and belongs to the technical field of molecular-assisted breeding of tetraploid long oysters. Background Art

[0002] The giant oyster (Crassostrea gigas) is the largest and most productive marine commercial shellfish aquacultured in my country and the world. It boasts high fatness, rapid growth, nutrient richness, and wide adaptability. Triploid giant oysters are more nutritious, and their low fertility ensures their gonads are plump year-round, meeting market demand year-round. The tetraploid giant oyster, however, is the core germplasm for producing fully triploid giant oysters. Advances in breeding technology have shifted the primary method for obtaining tetraploid giant oysters from traditional drug-induced breeding to self-propagation. This selective breeding has significantly accelerated the breeding of tetraploid oysters. Currently, family selection is a key breeding method for tetraploid giant oysters, enabling the generation of families with stable ploidy and excellent traits, facilitating the development of new varieties. However, family selection requires each family to be cultured individually, significantly increasing the labor and material resources required. Furthermore, family selection requires accurate documentation of the pedigree information of each family. Inaccurate pedigrees can reduce the accuracy of genetic evaluations, hindering germplasm improvement. Furthermore, incorrect pedigree information can increase the risk of inbreeding depression and lead to germplasm degradation. However, in large-scale aquaculture, pedigree information recording errors are common. Therefore, a method for efficiently and accurately recording pedigree relationships is needed to promote the stable development of tetraploid long oyster core germplasm breeding.

[0003] Parentage testing is a scientific method that uses genetic marker analysis technology to compare the similarity of genetic material between individuals and determine whether there is a biological relationship based on genetic laws. DNA molecular marker identification technology has become the preferred technology for parentage testing due to its high accuracy and specificity. This method determines parentage by detecting whether the inheritance pattern of DNA molecular markers between individuals conforms to Mendel's law. With the development of high-throughput sequencing, the DNA molecular markers used in parentage testing have evolved from the original restriction endonuclease length polymorphism (RFLP) to simple sequence repeats (SSR) and single nucleotide polymorphisms (SNP). Among them, compared with RFLP and SSR, SNP as a third-generation genetic marker has the following advantages: (1) It is widely distributed in the genome and has a large number; (2) The mutation and recombination rate is low and the repeatability is high; (3) The polymorphism is high; (4) The typing efficiency is high and the error rate is low. At the same time, studies have shown that a relatively small number of SNPs can accurately determine the parentage relationship between individuals. Based on the above advantages, SNP has become the main molecular marker for parentage testing.

[0004] Parentage determination software and R packages such as Cervus, PaternityT, and PediHaplotyper have been widely used in livestock and marine biology due to their high accuracy and efficiency. The Cervus software suite accurately identifies paternity between individuals by simulating non-paternity probabilities and calculating the likelihood ratio logarithm (LOD) between samples. PaternityT constructs a cumulative paternity index and a paternity probability matrix to calculate the cumulative non-paternity exclusion probability, enabling paternity determination. Both approaches have been widely used in marine biology. For example, Cervus accurately identifies kinship between channel catfish populations based on 116 SNPs. PaternityT accurately identifies paternity between individuals of yellow croaker by calculating the cumulative non-paternity exclusion probability for mixed samples from different families, accurately assigning them to family groups. However, these software packages only support analysis of diploid organisms and are not suitable for the more genetically complex tetraploid organisms. To date, there are no software or R packages specifically designed for paternity determination in tetraploid organisms. Therefore, it is necessary to combine the genetic laws of tetraploids, develop parentage identification methods for tetraploids, and improve the universality of parentage identification methods.

[0005] For the tetraploid long oyster, an important aquaculture species, the SNP site combination specifically used to efficiently and accurately identify its parent-offspring relationship has not yet been systematically screened and identified, and there is currently no method to identify its parent-offspring relationship. Summary of the Invention

[0006] In order to address the shortcomings of the existing technology, the purpose of the present invention is to screen out SNP sites that can efficiently and accurately identify the parent-offspring relationship of tetraploid long oysters, and based on this, develop a method suitable for identifying the parent-offspring relationship of tetraploid long oysters, aiming to apply this method in the tetraploid long oyster breeding industry to solve the problem that the parent-offspring relationship cannot be accurately recorded during the breeding process of tetraploid long oysters.

[0007] In order to achieve the above objectives, the present invention adopts the following technical solutions: A method for identifying parentage of tetraploid long oysters based on SNP loci comprises the following steps: (1) Sequencing the adductor muscle of the tetraploid oyster to be tested; (2) Compare the reference genome of Crassostrea gigas to obtain the genotypes of the designated SNP sites of the tetraploid Crassostrea gigas. The GenBank accession number of the reference genome of Crassostrea gigas is GCA_011032805.1. The genotypes of the designated SNP sites are as follows: Chromosome 1: G / A mutation at 121890bp, A / G mutation at 1057743bp, T / G mutation at 1282310bp, G / T mutation at 1672864bp, T / C mutation at 2377246bp, T / A mutation at 3067043bp, T / C mutation at 3586786bp, A / T mutation at 3999280bp, T / C mutation at 4728772bp, T / C mutation at 5044925bp, T / C mutation at 5300940bp, A / G mutation at 5594188bp, C / T mutation at 5903109bp, T / G mutation at 6330374bp, 6833 T / C mutation at 762bp, A / G mutation at 7943920bp, A / T mutation at 8487382bp, A / G mutation at 9704220bp, T / C mutation at 10224929bp, T / C mutation at 10862004bp, T / G mutation at 11750498bp, T / C mutation at 11990399bp, T / C mutation at 12218512bp, G / A mutation at 12483376bp, G / A mutation at 13361474bp, G / T mutation at 13765448bp, T / A mutation at 14300439bp, T / A mutation at 15205025bp, T / A mutation at 15645 A / G mutation at 080bp, G / T mutation at 15940394bp, T / C mutation at 17045095bp, T / C mutation at 17268995bp, T / A mutation at 17538044bp, A / T mutation at 17899956bp, G / T mutation at 18451368bp, C / T mutation at 18760584bp, C / G mutation at 19501089bp, G / T mutation at 21531956bp, T / A mutation at 22401721bp, T / A mutation at 24526392bp, A / T mutation at 25189059bp, G / A mutation at 25603252bp, A / T mutation at 161161bp, C / T mutation at 26611917bp, A / C mutation at 27004404bp, A / G mutation at 27425048bp, C / T mutation at 27805924bp, A / T mutation at 28632425bp, A / T mutation at 29021720bp, C / T mutation at 33066462bp, A / T mutation at 33718201bp, G / A mutation at 34717118bp, A / C mutation at 35189356bp, C / T mutation at 36608599bp, C / T mutation at 36886136bp, C / A mutation at 37186110bp,G / C mutation at 37483058bp, C / A mutation at 38812679bp, A / G mutation at 39546717bp, C / T mutation at 40317002bp, A / G mutation at 41392051bp, G / A mutation at 43046392bp, C / T mutation at 43389852bp, T / A mutation at 45510791bp, A / G mutation at 46464535bp, A / G mutation at 47037306bp, T / G mutation at 47275653bp, A / T mutation at 47543454bp, A / T mutation at 48596129bp, A / T mutation at 49583230bp / G mutation, T / C mutation at 49868879bp, T / G mutation at 50086266bp, T / G mutation at 50302873bp, A / C mutation at 51214188bp, C / T mutation at 51485365bp, G / A mutation at 52121167bp, A / T mutation at 52424658bp, C / T mutation at 52758029bp, A / G mutation at 53158981bp, C / T mutation at 53542635bp, T / C mutation at 53910234bp, C / T mutation at 54359105bp, T / C mutation at 55808097bp, bp T / A mutation, 57138543bp C / T mutation, 58299726bp G / A mutation, 59081390bp C / T mutation, 59774992bp G / T mutation, 60482812bp T / G mutation, 60889634bp C / T mutation, 61245346bp G / A mutation, 61505117bp C / G mutation, 61817752bp C / A mutation, 62517352bp A / T mutation, 62882914bp T / C mutation, 63105443bp A / G mutation, 64259558bp C / T mutation, 652 C / A mutation at 52225bp, T / A mutation at 66625596bp, A / G mutation at 66835219bp, C / T mutation at 67248469bp, T / A mutation at 67503687bp, A / G mutation at 67906236bp, C / T mutation at 68247170bp, C / T mutation at 68945233bp, T / C mutation at 69565892bp, G / A mutation at 70058662bp, A / G mutation at 70314116bp, A / T mutation at 70765241bp, A / G mutation at 72411555bp, T / C mutation at 73822739bp,A / G mutation at 74137180 bp, A / G mutation at 75053096 bp, A / C mutation at 75443458 bp, A / G mutation at 76271402 bp, T / A mutation at 76958049 bp, C / G mutation at 77190736 bp, T / A mutation at 77491425 bp, A / T mutation at 77966387 bp, C / T mutation at 78744623 bp; Chromosome 2: T / C mutation at 126208bp, G / A mutation at 530724bp, A / T mutation at 1030962bp, C / G mutation at 1883338bp, T / A mutation at 2385548bp, A / G mutation at 2612768bp, T / C mutation at 2905649bp, T / C mutation at 3408267bp, A / G mutation at 4209622bp, T / C mutation at 4735213bp, C / G mutation at 5034762bp, C / A mutation at 5605969bp, A / G mutation at 5935587bp, T / G mutation at 6189866bp, A / T mutation at 484bp, A / G mutation at 7186787bp, A / G mutation at 7919353bp, T / A mutation at 8150271bp, T / C mutation at 8400126bp, A / G mutation at 8695230bp, G / T mutation at 9416767bp, C / T mutation at 9823074bp, T / C mutation at 10397975bp, A / T mutation at 10863126bp, A / G mutation at 11074140bp, A / G mutation at 11296287bp, A / G mutation at 11502471bp, C / G mutation at 12364482bp, and 12574976 bp T / G mutation, T / C mutation at 12794499bp, C / A mutation at 13083777bp, T / C mutation at 13511367bp, T / C mutation at 13877414bp, C / A mutation at 14299527bp, G / A mutation at 14524308bp, A / G mutation at 14736903bp, G / A mutation at 15063383bp, A / G mutation at 15597292bp, T / A mutation at 15867029bp, G / A mutation at 16101395bp, C / A mutation at 16326428bp, T / C mutation at 16905050bp, and 172 A / T mutation at 66489bp, A / T mutation at 17579365bp, G / T mutation at 18124593bp, G / C mutation at 18474645bp, A / G mutation at 18705311bp, C / A mutation at 19267782bp, A / G mutation at 21377238bp, T / A mutation at 22776786bp, A / C mutation at 23709017bp, A / G mutation at 24156723bp, A / C mutation at 24530341bp, G / A mutation at 25094735bp, G / C mutation at 25421687bp, A / G mutation at 25763249bp,C / T mutation at 25980517bp, T / A mutation at 27508872bp, T / C mutation at 28212532bp, C / T mutation at 29113539bp, G / C mutation at 29485500bp, T / C mutation at 29809495bp, A / T mutation at 30098613bp, C / T mutation at 30391267bp, A / G mutation at 31955766bp, G / A mutation at 32672496bp, C / T mutation at 33448952bp, C / T mutation at 34565522bp, G / T mutation at 34854460bp, T at 35956877bp / G mutation, G / A mutation at 37299837bp, T / A mutation at 37768791bp, A / G mutation at 38364999bp, C / T mutation at 38772687bp, A / T mutation at 39119217bp, G / A mutation at 39791727bp, G / A mutation at 40956496bp, A / T mutation at 42059421bp, C / T mutation at 42389769bp, A / G mutation at 42831401bp, T / C mutation at 43232251bp, T / A mutation at 43943244bp, A / T mutation at 44224577bp, 44910891 bp A / T mutation, 46742409bp T / G mutation, 47231207bp A / G mutation, 47919343bp A / T mutation, 48360872bp G / A mutation, 49359594bp G / T mutation, 49898952bp G / A mutation, 50319548bp G / A mutation, 51049661bp T / C mutation, 51306897bp A / C mutation, 51516277bp A / G mutation, 52191511bp A / G mutation, 52579004bp G / A mutation, 53640899bp C / G mutation, 5397 A / T mutation at 6693bp, T / C mutation at 54891372bp, C / T mutation at 56410343bp, G / C mutation at 56902918bp, A / G mutation at 57143044bp, G / A mutation at 57422435bp, G / A mutation at 58005227bp, T / C mutation at 58329882bp, T / C mutation at 58802812bp, T / C mutation at 59036884bp, G / T mutation at 60176975bp, G / A mutation at 60593132bp, A / G mutation at 60929419bp, A / G mutation at 61642762bp;Chromosome 3: T / C mutation at 167511bp, G / T mutation at 444560bp, T / G mutation at 1740957bp, A / C mutation at 2247068bp, A / T mutation at 2663410bp, A / G mutation at 3153958bp, A / G mutation at 3443363bp, T / A mutation at 3918303bp, T / A mutation at 4164736bp, T / G mutation at 4395261bp, G / A mutation at 5114758bp, A / T mutation at 5385938bp, C / T mutation at 5778988bp, G / T mutation at 6150113bp, A / T mutation at 63bp, T / A mutation at 7403440bp, C / G mutation at 8943900bp, T / A mutation at 9256892bp, T / C mutation at 9919466bp, A / C mutation at 10174343bp, A / T mutation at 11633388bp, G / A mutation at 12017984bp, A / G mutation at 12915914bp, C / T mutation at 14184322bp, A / T mutation at 14610187bp, C / T mutation at 15112027bp, A / C mutation at 15742955bp, T / C mutation at 18055450bp, 05bp T / C mutation, A / T mutation at 20806332bp, C / T mutation at 21239659bp, A / T mutation at 21451371bp, A / G mutation at 21908751bp, C / T mutation at 22900014bp, T / G mutation at 23437899bp, C / G mutation at 24068761bp, C / T mutation at 24297401bp, A / G mutation at 25467666bp, A / T mutation at 26854208bp, G / A mutation at 27503027bp, C / T mutation at 27774729bp, T / C mutation at 28167758bp, A / G mutation at 499237bp, G / A mutation at 29844353bp, A / G mutation at 30081511bp, G / T mutation at 30294248bp, G / A mutation at 31271920bp, C / T mutation at 31611643bp, C / T mutation at 31900387bp, A / T mutation at 32420847bp, A / G mutation at 33314059bp, C / G mutation at 33562476bp, A / G mutation at 34537410bp, A / C mutation at 35525002bp, G / A mutation at 36168038bp, G / A mutation at 37291953bp,G / C mutation at 37938922bp, T / C mutation at 38158633bp, G / T mutation at 38470830bp, G / A mutation at 39158628bp, G / A mutation at 39719498bp, A / C mutation at 40603131bp, A / C mutation at 40865370bp, T / G mutation at 41259567bp, T / C mutation at 41597965bp, G / A mutation at 41910605bp, T / A mutation at 42938999bp, A / T mutation at 43500081bp, C / T mutation at 44213296bp, A / G mutation at 44459501bp, C / A mutation at 45077111bp, A / T mutation at 46562102bp, T / A mutation at 47015472bp, A / C mutation at 47248636bp, A / G mutation at 47755545bp, G / A mutation at 48068295bp, G / A mutation at 48403665bp, C / A mutation at 48859386bp, A / G mutation at 49155040bp, A / T mutation at 49514429bp, T / A mutation at 49801952bp, G / C mutation at 50508624bp, T / C mutation at 50867456bp, C / T mutation at 51172288bp, G / A mutation at 51555200bp, T / A mutation at 51897141bp, C / G mutation at 53046160bp, T / A mutation at 53359071bp, T / A mutation at 53636779bp, C / T mutation at 53869457bp, T / C mutation at 54594267bp, T / C mutation at 55269130bp, A / G mutation at 55572008bp, A / C mutation at 55896201bp, C / T mutation at 56783882bp, A / G mutation at 57104330 bp, C / G mutation at 57532214 bp, T / C mutation at 57807917 bp, T / A mutation at 58451281 bp, C / G mutation at 58805029 bp, A / C mutation at 59273801 bp, C / A mutation at 59815549 bp, A / G mutation at 60128400 bp, G / A mutation at 60681360 bp, C / T mutation at 61275765 bp, C / T mutation at 61522130 bp, C / A mutation at 61801828 bp, T / C mutation at 62069187 bp; Chromosome 4: C / A mutation at 489947bp, G / C mutation at 886286bp, A / G mutation at 2582998bp, G / A mutation at 3139977bp, G / A mutation at 3595215bp, T / A mutation at 3859888bp, A / G mutation at 4344155bp, C / A mutation at 4566835bp, A / G mutation at 5145569bp, T / C mutation at 6456871bp, G / C mutation at 7321215bp, T / A mutation at 7817588bp, T / G mutation at 8179181bp, G / T mutation at 8910032bp, 6bp G / T mutation, T / G mutation at 9552736bp, G / A mutation at 10022246bp, T / G mutation at 10354612bp, G / A mutation at 10907817bp, C / T mutation at 11734493bp, G / A mutation at 12580554bp, T / C mutation at 13260828bp, T / C mutation at 13765169bp, C / T mutation at 14310601bp, T / C mutation at 16355043bp, G / A mutation at 17630046bp, T / A mutation at 19429489bp, A / T mutation at 19679728bp, and 21079 A / G mutation at 213bp, T / C mutation at 22164490bp, A / C mutation at 22482582bp, T / A mutation at 22780082bp, C / T mutation at 23190102bp, A / G mutation at 23587372bp, A / G mutation at 24228102bp, A / G mutation at 24961164bp, G / T mutation at 25252848bp, A / T mutation at 25844871bp, G / T mutation at 26112318bp, G / A mutation at 27430034bp, C / A mutation at 27889194bp, T / C mutation at 28186129bp, G / T mutation at 862600bp, T / A mutation at 29613029bp, A / T mutation at 29849521bp, C / T mutation at 30074147bp, G / T mutation at 30551177bp, A / C mutation at 31624189bp, T / C mutation at 31868549bp, T / C mutation at 32085476bp, T / C mutation at 33390588bp, T / C mutation at 33885908bp, T / A mutation at 34321828bp, C / A mutation at 34763173bp, A / G mutation at 36691587bp, A / G mutation at 36995284bp,A / C mutation at 37364904bp, C / T mutation at 39303404bp, G / C mutation at 40040754bp, T / C mutation at 40300785bp, T / C mutation at 41007997bp, T / C mutation at 41213469bp, A / G mutation at 41586776bp, A / G mutation at 42194520bp, G / T mutation at 42525010bp, C / T mutation at 42726376bp, T / G mutation at 43723088bp, A / T mutation at 44141447bp, T / C mutation at 44798262bp, A / G mutation at 45230255bp, A / G mutation at 45691471bp, T / A mutation at 45924080bp, A / G mutation at 46335423bp, G / A mutation at 46609807bp, C / T mutation at 47649738bp, C / A mutation at 48054473bp, G / A mutation at 49037444bp, T / C mutation at 49356447bp, T / A mutation at 49823141bp, C / T mutation at 50253140bp, T / A mutation at 50495698bp, G / A mutation at 50782734bp, G / A mutation at 51328829bp, T / C mutation at 51912827bp, T / C mutation at 52524776bp, C / T mutation at 52849059bp, C / T mutation at 53628954bp, C / T mutation at 54121390bp, C / T mutation at 54500655bp, A / T mutation at 54780363bp, C / A mutation at 55043635bp, T / C mutation at 55269248bp, C / T mutation at 56009969bp, T / C mutation at 56360644bp, T / A mutation at 56621645bp, A / G mutation at 57033970 bp, T / A mutation at 57487468 bp, C / T mutation at 58248810 bp, A / G mutation at 58569969 bp, C / A mutation at 58804428 bp, G / A mutation at 59177752 bp, A / G mutation at 59780097 bp, C / A mutation at 60118503 bp, G / T mutation at 60642124 bp, T / A mutation at 60872702 bp, T / G mutation at 61335564 bp, G / A mutation at 61988945 bp, G / T mutation at 62219348 bp; Chromosome 5: C / G mutation at 879645bp, G / A mutation at 1415586bp, C / T mutation at 2111293bp, C / A mutation at 2314335bp, A / T mutation at 2635565bp, G / A mutation at 2920345bp, T / C mutation at 3467779bp, T / A mutation at 3884050bp, C / G mutation at 4393916bp, A / C mutation at 4660080bp, T / C mutation at 5064983bp, G / A mutation at 5762044bp, G / T mutation at 6544275bp, G / T mutation at 6749408bp, G / T mutation at 699 A / G mutation at 7717bp, T / G mutation at 7345402bp, A / G mutation at 7654532bp, A / G mutation at 7932212bp, A / C mutation at 8303258bp, T / C mutation at 8661151bp, T / C mutation at 9176468bp, C / A mutation at 9464239bp, T / C mutation at 9931584bp, A / G mutation at 10539507bp, A / T mutation at 10875599bp, G / A mutation at 11217368bp, C / T mutation at 11702291bp, T / G mutation at 12003330bp, and 12366955 bp A / T mutation, 12808603bp T / C mutation, 13875823bp T / C mutation, 14157737bp A / G mutation, 15150248bp G / A mutation, 15517737bp A / G mutation, 15749958bp A / G mutation, 16257325bp A / G mutation, 16594460bp T / C mutation, 17783174bp T / A mutation, 18355754bp C / A mutation, 18594492bp G / A mutation, 18998718bp C / T mutation, 202 T / C mutation at 53708bp, A / C mutation at 20548723bp, G / A mutation at 22378076bp, A / G mutation at 23213346bp, A / C mutation at 23730799bp, T / C mutation at 23996432bp, C / T mutation at 24241420bp, G / A mutation at 24721774bp, T / C mutation at 26586598bp, A / T mutation at 26893151bp, C / T mutation at 27345167bp, T / C mutation at 27679094bp, C / G mutation at 28185342bp, T / A mutation at 29149844bp,C / A mutation at 29649330bp, G / A mutation at 30587198bp, C / A mutation at 30997742bp, G / T mutation at 31256011bp, T / C mutation at 31705376bp, T / C mutation at 32770700bp, G / C mutation at 33339763bp, A / G mutation at 33573910bp, C / T mutation at 34514253bp, A / G mutation at 35112239bp, T / C mutation at 35669268bp, T / A mutation at 36705272bp, C / T mutation at 36922640bp, A / C mutation at 37131668bp / G mutation, C / T mutation at 37581075bp, C / T mutation at 38542724bp, G / A mutation at 39044859bp, G / C mutation at 39688041bp, G / A mutation at 40000656bp, T / C mutation at 40395464bp, G / A mutation at 41001494bp, T / C mutation at 41681731bp, T / C mutation at 42174184bp, A / T mutation at 42576737bp, A / G mutation at 43098562bp, A / G mutation at 43487720bp, A / G mutation at 43692812bp, 44169923 bp A / C mutation, 44373192bp A / C mutation, 44895351bp C / T mutation, 45190288bp A / G mutation, 46681737bp T / A mutation, 47488756bp T / C mutation, 49326052bp T / C mutation, 49639111bp G / T mutation, 50108002bp C / G mutation, 50436257bp T / A mutation, 50696264bp A / T mutation, 50903386bp T / C mutation, 51419745bp A / G mutation, 51669577bp C / T mutation, 524 T / A mutation at 14271bp, A / C mutation at 53368505bp, A / G mutation at 53605312bp, T / C mutation at 53894100bp, T / A mutation at 54916254bp, A / G mutation at 55477238bp, A / C mutation at 55999091bp, G / C mutation at 56214132bp, G / A mutation at 57070120bp, T / A mutation at 57313644bp, A / C mutation at 57666549bp, G / T mutation at 57877313bp, C / T mutation at 58133420bp, G / A mutation at 58891833bp,T / C mutation at 59245611bp, A / T mutation at 59751484bp, A / G mutation at 59976681bp; Chromosome 6: G / A mutation at 587350bp, A / G mutation at 1075497bp, T / A mutation at 1679222bp, G / T mutation at 2172358bp, T / C mutation at 2739350bp, G / A mutation at 3206148bp, C / A mutation at 3421291bp, G / A mutation at 3843416bp, A / T mutation at 4064437bp, G / T mutation at 4469234bp, T / C mutation at 4760643bp, T / A mutation at 4992337bp, A / T mutation at 5778802bp, A / C mutation at 6400 T / C mutation at 355bp, G / A mutation at 6932249bp, G / A mutation at 8026496bp, C / T mutation at 8885122bp, T / C mutation at 9620041bp, G / T mutation at 10908040bp, C / T mutation at 11117547bp, A / G mutation at 12753363bp, C / T mutation at 14268050bp, T / G mutation at 14586618bp, G / C mutation at 14879882bp, G / A mutation at 15232542bp, A / C mutation at 15807647bp, C / T mutation at 16152642bp, and 164216 A / G mutation at 43bp, A / T mutation at 16738467bp, C / A mutation at 17154757bp, G / A mutation at 17669050bp, T / A mutation at 17995298bp, A / T mutation at 18653957bp, T / C mutation at 18914180bp, T / A mutation at 19545790bp, C / T mutation at 20589164bp, A / G mutation at 21516044bp, T / A mutation at 22028166bp, A / T mutation at 22624328bp, T / A mutation at 23340429bp, C / T mutation at 23832772bp, A / G mutation at 266242bp, A / C mutation at 25073811bp, A / G mutation at 26222500bp, A / C mutation at 26774119bp, T / C mutation at 27210621bp, C / T mutation at 27689291bp, G / A mutation at 28235309bp, T / C mutation at 28437764bp, C / T mutation at 28668444bp, T / A mutation at 29329850bp, A / G mutation at 29645718bp, G / T mutation at 30524553bp, G / A mutation at 31121205bp, C / T mutation at 31359984bp,A / G mutation at 32106045bp, G / A mutation at 32457595bp, T / C mutation at 32753400bp, T / A mutation at 33117124bp, T / C mutation at 33570502bp, T / G mutation at 34176629bp, C / T mutation at 34667060bp, A / G mutation at 38181718bp, T / A mutation at 39546371bp, G / C mutation at 39866670bp, G / T mutation at 40351063bp, A / G mutation at 40681703bp, T / C mutation at 41023800bp, G / T mutation at 41459997bp, T / C mutation at 42016811bp, T / C mutation at 42364688bp, T / A mutation at 42605575bp, C / A mutation at 43099589bp, C / T mutation at 43383766bp, C / T mutation at 43791635bp, A / T mutation at 44792472bp, C / T mutation at 45098188bp, A / G mutation at 45791769bp, A / G mutation at 46506334bp, T / C mutation at 47036051bp, G / C mutation at 47496058bp, C / T mutation at 47812848bp, A / C mutation at 48271374bp, T / A mutation at 49169061bp, A / T mutation at 49429004bp, C / T mutation at 49743523 bp, T / C mutation at 50128359 bp, A / G mutation at 50887722 bp, A / T mutation at 51813382 bp, C / T mutation at 52554583 bp, T / C mutation at 53230845 bp, C / T mutation at 54083112 bp, C / T mutation at 54380581 bp, C / T mutation at 55428291 bp, A / C mutation at 56179611 bp; Chromosome 7: A / G mutation at 603788bp, G / T mutation at 896391bp, T / C mutation at 1282724bp, G / T mutation at 1535834bp, G / A mutation at 1795922bp, T / A mutation at 2124131bp, G / T mutation at 2694802bp, G / A mutation at 3023240bp, T / C mutation at 3227288bp, T / C mutation at 3539581bp, T / C mutation at 3946542bp, T / A mutation at 4409015bp, G / A mutation at 4610418bp, A / G mutation at 5099145bp, 5726 T / C mutation at 774bp, T / C mutation at 6090181bp, T / A mutation at 7174959bp, G / T mutation at 7732660bp, T / A mutation at 7943436bp, C / T mutation at 8179975bp, A / T mutation at 9080480bp, T / A mutation at 9322949bp, A / G mutation at 9569794bp, T / C mutation at 9911214bp, A / C mutation at 10663904bp, T / G mutation at 10877221bp, G / A mutation at 11467271bp, T / A mutation at 11736967bp, and 12053878b p, T / C mutation at 13007410bp, C / T mutation at 13768058bp, A / G mutation at 15198787bp, G / A mutation at 15771972bp, T / G mutation at 16027470bp, G / A mutation at 16465374bp, T / G mutation at 16680508bp, A / G mutation at 17192258bp, T / C mutation at 17558072bp, C / T mutation at 17969940bp, C / T mutation at 18480038bp, C / T mutation at 18732421bp, A / C mutation at 19150831bp, A / T mutation at 8092bp, C / T mutation at 19653197bp, T / C mutation at 20077315bp, T / G mutation at 20839591bp, A / T mutation at 21779850bp, A / G mutation at 23771667bp, C / A mutation at 24586945bp, A / G mutation at 25311728bp, A / C mutation at 25544379bp, T / C mutation at 26417838bp, G / C mutation at 26642847bp, A / G mutation at 26965429bp, A / T mutation at 29758485bp, T / A mutation at 30141591bp,C / T mutation at position 30468300 bp, T / C mutation at position 31400778 bp, G / T mutation at position 31778180 bp, A / G mutation at position 32075251 bp, C / T mutation at position 33230067 bp, C / T mutation at position 33744108 bp, T / A mutation at position 36023112 bp, T / A mutation at position 36396469 bp, A / G mutation at position 36693769 bp, A / T mutation at position 36913195 bp, C / T mutation at position 37760599 bp, T / A mutation at position 38161672 bp, G / T mutation at position 38702757 bp, A / G mutation at position 39210903 bp, A / T mutation at position 39699534 bp, G / C mutation at position 40385742 bp, T / C mutation at position 41026442 bp, C / A mutation at position 41307086 bp, G / A mutation at position 42969755 bp, T / C mutation at position 43571847 bp, A / G mutation at position 43993986 bp, T / C mutation at position 44394370 bp, G / A mutation at position 44703340 bp, A / G mutation at position 45524469 bp, C / T mutation at position 46045824 bp, A / G mutation at position 47359820 bp, C / A mutation at position 47598126 bp, A / G mutation at position 49292205 bp, T / C mutation at position 50492560 bp, T / C mutation at position 50866106 bp, C / T mutation at position 51554060 bp, T / G mutation at position 51867069 bp, T / C mutation at position 52186344 bp, A / G mutation at position 52386979 bp, A / G mutation at position 52719201 bp, C / T mutation at position 53780380 bp, G / A mutation at position 54228211 bp, A / C mutation at position 54901003 bp;, Chromosome 8: G / A mutation at 412562bp, T / A mutation at 985611bp, T / C mutation at 1303653bp, C / T mutation at 1544349bp, T / C mutation at 1922320bp, T / C mutation at 2361255bp, C / A mutation at 2719405bp, A / G mutation at 3000604bp, T / C mutation at 3288220bp, G / C mutation at 3518544bp, T / C mutation at 3933031bp, T / C mutation at 4572621bp, T / C mutation at 5346224bp, A / T mutation at 5602024bp, The mutations at 95bp were T / C, 6172964bp C / T, 6581392bp T / A, 7427921bp T / A, 7653299bp T / C, 10833172bp G / A, 11143937bp C / T, 11563545bp C / T, 12240151bp T / C, 12664163bp A / G, 12976720bp C / A, 13428217bp C / G, 13690299bp T / A, 14184965bp C / A, 145407 A / G mutation at 137bp, A / C mutation at 14927534bp, T / A mutation at 15164050bp, A / C mutation at 16228337bp, C / G mutation at 16839194bp, T / A mutation at 17073823bp, A / G mutation at 17560361bp, C / T mutation at 17805117bp, C / T mutation at 18668691bp, C / T mutation at 19077073bp, A / T mutation at 19321381bp, A / G mutation at 20212862bp, A / G mutation at 20498190bp, T / C mutation at 20748906bp, A / G mutation at 088463bp, A / G mutation at 21591018bp, A / G mutation at 22165379bp, A / T mutation at 23067147bp, T / C mutation at 23844408bp, A / G mutation at 24104507bp, T / A mutation at 24665627bp, G / A mutation at 25079866bp, T / G mutation at 25958988bp, A / G mutation at 27292396bp, G / A mutation at 27511452bp, C / T mutation at 27712853bp, G / A mutation at 27981923bp, G / A mutation at 28485338bp,G / A mutation at 28816819bp, C / A mutation at 29401272bp, C / T mutation at 29733986bp, C / T mutation at 30150021bp, A / T mutation at 30361015bp, A / C mutation at 30665469bp, T / G mutation at 30927726bp, C / G mutation at 31612011bp, and C / T mutation at 31949923 bp C / G mutation, 32350947bp T / C mutation, 32829805bp T / C mutation, 33830538bp T / C mutation, 34428621bp T / C mutation, 34631910bp G / A mutation, 34878333bp C / A mutation, 35167435bp T / C mutation, 35384611bp G / A mutation, C / T mutation at 36295451bp, A / G mutation at 36758686bp, T / C mutation at 38408273bp, T / C mutation at 39084417bp, T / C mutation at 40515577bp, A / G mutation at 40784564bp, C / T mutation at 41770053bp, G / T mutation at 42205041bp, and bp G / A mutation, G / A mutation at 43574397bp, T / G mutation at 44913196bp, C / A mutation at 46413323bp, C / T mutation at 47254739bp, T / C mutation at 47569565bp, C / T mutation at 48443108bp, A / T mutation at 48947202bp, T / C mutation at 50071965bp; Chromosome 9: T / C mutation at 217853bp, A / G mutation at 583042bp, A / C mutation at 796208bp, T / C mutation at 1046351bp, T / G mutation at 1327845bp, G / A mutation at 2838975bp, A / G mutation at 3240786bp, T / C mutation at 3675421bp, G / T mutation at 4651246bp, C / T mutation at 4959856bp, C / T mutation at 5227751bp, C / T mutation at 5933068bp, G / A mutation at 6728623bp, C / T mutation at 7235075bp, 74929 C / G mutation at 85bp, A / T mutation at 7915378bp, A / G mutation at 8155620bp, A / C mutation at 8437140bp, C / A mutation at 9041049bp, T / G mutation at 9320125bp, G / C mutation at 9523788bp, A / G mutation at 10256842bp, A / T mutation at 10516635bp, C / G mutation at 11295005bp, C / G mutation at 11568023bp, T / A mutation at 12159413bp, C / T mutation at 12529731bp, T / G mutation at 12735255bp, and 13042605 bp C / A mutation, A / G mutation at 13493638bp, A / T mutation at 13853217bp, G / A mutation at 14058546bp, A / T mutation at 15219775bp, G / T mutation at 15704220bp, G / A mutation at 16140181bp, T / C mutation at 16412881bp, T / C mutation at 17010061bp, T / C mutation at 17928904bp, C / T mutation at 18193136bp, T / C mutation at 18452448bp, T / C mutation at 18653747bp, A / T mutation at 18878597bp, 196 C / T mutation at 85478bp, G / A mutation at 20221044bp, T / C mutation at 20993779bp, G / T mutation at 21586001bp, A / G mutation at 22002234bp, C / T mutation at 22394787bp, T / A mutation at 22969695bp, A / T mutation at 23876847bp, C / T mutation at 25194999bp, A / G mutation at 25462264bp, A / T mutation at 25741402bp, A / G mutation at 26959004bp, C / A mutation at 27955752bp, A / G mutation at 28258725bp,A / T mutation at position 29236123 bp, A / G mutation at position 29908750 bp, C / T mutation at position 30127537 bp, T / C mutation at position 31200772 bp, C / T mutation at position 31445189 bp, T / C mutation at position 31759757 bp, C / A mutation at position 32418276 bp, T / C mutation at position 33430899 bp, G / A mutation at position 33857260 bp, A / G mutation at position 34836159 bp, A / G mutation at position 35910999 bp, T / C mutation at position 36657698 bp, A / T mutation at position 37111454 bp, A / T mutation at position 38333366 bp, G / T mutation at position 38956659 bp, T / G mutation at position 41437696 bp, G / T mutation at position 41655725 bp, T / C mutation at position 42066006 bp, G / C mutation at position 43778385 bp, T / C mutation at position 44072139 bp, C / T mutation at position 44341586 bp, T / C mutation at position 46347906 bp, C / A mutation at position 46681275 bp, A / G mutation at position 47089848 bp, T / C mutation at position 47332573 bp, G / A mutation at position 47545232 bp, T / C mutation at position 47787454 bp, T / A mutation at position 48000048 bp, T / C mutation at position 48209798 bp, T / C mutation at position 49478480 bp, G / A mutation at position 50204065 bp, C / T mutation at position 50425019 bp, T / G mutation at position 52132614 bp, T / C mutation at position 52785787 bp, C / T mutation at position 53663195 bp, C / T mutation at position 54046362 bp, T / G mutation at position 54605490 bp, T / C mutation at position 54826243 bp, Chromosome 10: T / C mutation at 1534163bp, T / G mutation at 2455069bp, C / T mutation at 3038564bp, T / C mutation at 4197932bp, A / T mutation at 4800809bp, A / T mutation at 5676331bp, T / C mutation at 6642222bp, A / G mutation at 7400941bp, T / C mutation at 7668457bp, T / C mutation at 7897420bp, A / C mutation at 8180708bp, C / T mutation at 8755317bp, G / A mutation at 9215005bp, C / A mutation at 9779335bp, 1040 A / G mutation at 5131bp, A / G mutation at 10801950bp, T / C mutation at 11089097bp, C / A mutation at 11732224bp, T / C mutation at 11959146bp, C / T mutation at 12182036bp, A / G mutation at 12384483bp, T / A mutation at 12977779bp, C / G mutation at 13228019bp, G / A mutation at 13725920bp, A / G mutation at 14297949bp, C / T mutation at 14599462bp, A / C mutation at 15316648bp, C / T mutation at 16226209bp, T / A mutation at 125566bp, G / T mutation at 19323146bp, C / A mutation at 20329041bp, C / A mutation at 20545842bp, A / G mutation at 20831667bp, A / G mutation at 21077840bp, T / C mutation at 21855677bp, C / T mutation at 22076843bp, T / G mutation at 22285822bp, C / A mutation at 22573506bp, T / C mutation at 23139968bp, A / T mutation at 23927295bp, C / T mutation at 24217188bp, T / A mutation at 24958349bp, A / T mutation at 26265143bp, A / C mutation at 26789789bp, A / G mutation at 27013959bp, C / T mutation at 27270677bp, T / A mutation at 27522828bp, A / T mutation at 28230594bp, A / T mutation at 28621223bp, T / C mutation at 28976931bp, G / C mutation at 30161642bp, G / T mutation at 30602940bp, A / G mutation at 30978727bp, T / A mutation at 31649003bp, G / A mutation at 32503776bp, T / C mutation at 33082095bp,G / A mutation at 33398327bp, A / T mutation at 34352656bp, C / T mutation at 34825626bp, A / G mutation at 35322952bp, A / T mutation at 35627155bp, C / T mutation at 35967237bp, C / G mutation at 36252688bp, G / A mutation at 36722144bp, T / C mutation at 37789019bp; (3) Calculate the IBS and LOD of all tested parents and offspring based on the SNP typing results. The calculation formula is as follows: ,

[0008] Where n represents the number of successfully typed SNP sites, k represents the kth SNP site, G ik is the genotype of individual i at the kth SNP site, G jk is the genotype of individual j at the kth SNP site, I k (G ik , G jk ) represents the genotype matching score of individual i and individual j at the kth SNP site. When the genotypes of individual i and individual j at the kth SNP site can be matched as parent-offspring, I k (G ik , G jk ) is 1, otherwise I k (G ik , G jk ) is 0, P1 represents the probability that the candidate parent and the simulated parent produce offspring, and P2 represents the background probability; (4) Combining IBS and LOD to identify the parent-offspring relationship;

[0009] Preferably, in step (3), the probability P1 of the candidate parent and the simulated parent producing offspring is calculated as follows:

[0010] in, represents the probability of the candidate parent gametes appearing, represents the probability of the simulated parent gametes appearing, w represents the frequency of the simulated parent genotype in the population, m represents the number of mismatched genotypes between parents and offspring, and λ represents the mismatch penalty weight, which is 0.1.

[0011] Preferably, in step (3), when calculating the background probability P2, the genotypes of the simulated parents are determined based on the population allele frequencies.

[0012] Preferably, in step (4), the method for identifying the parent-offspring relationship of the parent to be tested and the offspring by combining IBS and LOD is specifically as follows: (1) When LOD is greater than 0, the candidate parent is likely to be the true parent, and at least one parent with the highest LOD is most likely to be the true parent of the offspring. At this time, the IBS between the candidate parent and the offspring should be higher than the preset threshold of 0.98; (2) When LOD is less than 0, the candidate parent cannot be the true parent.

[0013] The present invention is beneficial in that: (1) This invention discloses for the first time 1,000 SNP sites for parentage identification of tetraploid long oysters, providing data support for parentage identification of tetraploid long oysters and laying the foundation for the development of liquid phase chips for parentage identification of tetraploid long oysters, which has good application prospects. (2) Based on the tetraploid genetic law, the present invention has developed a method for identifying the parentage of tetraploid long oysters. Based on the two developed indicators, IBS and LOD, the parentage can be accurately identified through low-density SNP site information, which makes up for the current lack of molecular means for identifying the parentage of tetraploid long oysters. It also provides a guarantee for identifying and correcting potential pedigree errors, thereby improving the accuracy of molecular breeding of tetraploid long oysters.

[0014] (3) The parent-child relationship identification method provided by the present invention is low-cost and can be used for large-scale parent-child relationship determination and family map construction.

[0015] (4) The parentage identification method provided by the present invention is also applicable to the parentage identification of other tetraploid organisms and has high universality. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 It is the distribution diagram of LOD between samples with parent-child relationship; Figure 2 This is the LOD and IBS heat map between some samples. DETAILED DESCRIPTION

[0017] The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments.

[0018] 1. Screening for SNPs that can efficiently and accurately identify parentage in tetraploid oysters The method of the present invention for screening SNP sites that can efficiently and accurately identify the parentage relationship of tetraploid long oysters is as follows: 1. DNA purification, sequencing library construction and high-throughput sequencing were performed on the adductor muscle of tetraploid long oysters with clear parentage. Five individuals were collected from each of four tetraploid long oyster families, and each family included two parents and three offspring.

[0019] Adductor muscles of the tetraploid long oysters were collected and DNA was purified using the GenoPrep Polysaccharide and Polyphenol DNA Extraction Kit (Shijiazhuang Boredi Biotechnology Co., Ltd.). DNA concentration was determined using a Qubit 2.0 fluorescence quantification instrument, and DNA quality was assessed by 1% agarose gel electrophoresis.

[0020] Sequencing libraries were constructed using the GenoBaits® DNA Library Prep Kit for ILM.

[0021] The DNA fragments in the sequencing library were subjected to high-throughput sequencing using an MGISEQ-2000 sequencer (Shenzhen MGI Intelligent Manufacturing Technology Co., Ltd.) in PE150 mode.

[0022] 2. Genotyping of sequencing data The raw data were aligned to the long oyster reference genome (GenBank accession number: GCA_011032805.1), and genotyping was performed on the aligned data using GATK.

[0023] 3. Quality control of SNP sites Quality control was performed on the sites obtained by typing. First, indel sites and multi-allelic SNP sites were filtered out, and then the remaining di-allelic SNP sites were filtered according to the following criteria: (1) minor allele frequency (MAF) <0.35; (2) site detection rate <100%; (3) sequencing depth (DP) <100.

[0024] 4. Calculate the MAF, expected heterozygosity (Hexp), and polymorphism information content (PIC) of SNP sites that meet quality control standards, and screen SNP sites with high MAF, PIC, and Hepx from each chromosome. The number of SNP sites on each chromosome is positively correlated with the length of each chromosome, and the distance between adjacent SNP sites exceeds 200K.

[0025] The present invention ultimately screened out 1,000 SNP loci on 10 chromosomes for parentage identification in tetraploid long oysters. The information of these 1,000 SNP loci is as follows: Chromosome 1: G / A mutation at 121890bp, A / G mutation at 1057743bp, T / G mutation at 1282310bp, G / T mutation at 1672864bp, T / C mutation at 2377246bp, T / A mutation at 3067043bp, T / C mutation at 3586786bp, A / T mutation at 3999280bp, T / C mutation at 4728772bp, T / C mutation at 5044925bp, T / C mutation at 5300940bp, A / G mutation at 5594188bp, C / T mutation at 5903109bp, T / G mutation at 6330374bp, 6833 T / C mutation at 762bp, A / G mutation at 7943920bp, A / T mutation at 8487382bp, A / G mutation at 9704220bp, T / C mutation at 10224929bp, T / C mutation at 10862004bp, T / G mutation at 11750498bp, T / C mutation at 11990399bp, T / C mutation at 12218512bp, G / A mutation at 12483376bp, G / A mutation at 13361474bp, G / T mutation at 13765448bp, T / A mutation at 14300439bp, T / A mutation at 15205025bp, T / A mutation at 15645 A / G mutation at 080bp, G / T mutation at 15940394bp, T / C mutation at 17045095bp, T / C mutation at 17268995bp, T / A mutation at 17538044bp, A / T mutation at 17899956bp, G / T mutation at 18451368bp, C / T mutation at 18760584bp, C / G mutation at 19501089bp, G / T mutation at 21531956bp, T / A mutation at 22401721bp, T / A mutation at 24526392bp, A / T mutation at 25189059bp, G / A mutation at 25603252bp, A / T mutation at 161161bp, C / T mutation at 26611917bp, A / C mutation at 27004404bp, A / G mutation at 27425048bp, C / T mutation at 27805924bp, A / T mutation at 28632425bp, A / T mutation at 29021720bp, C / T mutation at 33066462bp, A / T mutation at 33718201bp, G / A mutation at 34717118bp, A / C mutation at 35189356bp, C / T mutation at 36608599bp, C / T mutation at 36886136bp, C / A mutation at 37186110bp,G / C mutation at 37483058bp, C / A mutation at 38812679bp, A / G mutation at 39546717bp, C / T mutation at 40317002bp, A / G mutation at 41392051bp, G / A mutation at 43046392bp, C / T mutation at 43389852bp, T / A mutation at 45510791bp, A / G mutation at 46464535bp, A / G mutation at 47037306bp, T / G mutation at 47275653bp, A / T mutation at 47543454bp, A / T mutation at 48596129bp, A / T mutation at 49583230bp / G mutation, T / C mutation at 49868879bp, T / G mutation at 50086266bp, T / G mutation at 50302873bp, A / C mutation at 51214188bp, C / T mutation at 51485365bp, G / A mutation at 52121167bp, A / T mutation at 52424658bp, C / T mutation at 52758029bp, A / G mutation at 53158981bp, C / T mutation at 53542635bp, T / C mutation at 53910234bp, C / T mutation at 54359105bp, T / C mutation at 55808097bp, bp T / A mutation, 57138543bp C / T mutation, 58299726bp G / A mutation, 59081390bp C / T mutation, 59774992bp G / T mutation, 60482812bp T / G mutation, 60889634bp C / T mutation, 61245346bp G / A mutation, 61505117bp C / G mutation, 61817752bp C / A mutation, 62517352bp A / T mutation, 62882914bp T / C mutation, 63105443bp A / G mutation, 64259558bp C / T mutation, 652 C / A mutation at 52225bp, T / A mutation at 66625596bp, A / G mutation at 66835219bp, C / T mutation at 67248469bp, T / A mutation at 67503687bp, A / G mutation at 67906236bp, C / T mutation at 68247170bp, C / T mutation at 68945233bp, T / C mutation at 69565892bp, G / A mutation at 70058662bp, A / G mutation at 70314116bp, A / T mutation at 70765241bp, A / G mutation at 72411555bp, T / C mutation at 73822739bp,A / G mutation at 74137180 bp, A / G mutation at 75053096 bp, A / C mutation at 75443458 bp, A / G mutation at 76271402 bp, T / A mutation at 76958049 bp, C / G mutation at 77190736 bp, T / A mutation at 77491425 bp, A / T mutation at 77966387 bp, C / T mutation at 78744623 bp; 2: T / C at position 126208, G / A at position 530724, A / T at position 1030962, C / G at position 1883338, T / A at position 2385548, A / G at position 2612768, T / C at position 2905649, T / C at position 3408267, A / G at position 4209622, T / C at position 4735213, C / G at position 5034762, C / A at position 5605969, A / G at position 5935587, T / G at position 6189866, A / T at position 6451484, A / G at position 7186787, A / G at position 7919353, T / A at position 8150271, T / C at position 8400126, A / G at position 8695230, G / T at position 9416767, C / T at position 9823074, T / C at position 10397975, A / T at position 10863126, A / G at position 11074140, A / G at position 11296287, A / G at position 11502471, C / G at position 12364482, T / G at position 12574976, T / C at position 12794499, C / A at position 13083777, T / C at position 13511367, T / C at position 13877414, C / A at position 14299527, G / A at position 14524308, A / G at position 14736903, G / A at position 15063383, A / G at position 15597292, T / A at position 15867029, G / A at position 16101395, C / A at position 16326428, T / C at position 16905050, A / T at position 17266489, A / T at position 17579365, G / T at position 18124593, G / C at position 18474645, A / G at position 18705311, C / A at position 19267782, A / G at position 21377238, T / A at position 22776786, A / C at position 23709017, A / G at position 24156723, A / C at position 24530341, G / A at position 25094735, G / C at position 25421687, A / G at position 25763249,C / T mutation at position 25980517 bp, T / A mutation at position 27508872 bp, T / C mutation at position 28212532 bp, C / T mutation at position 29113539 bp, G / C mutation at position 29485500 bp, T / C mutation at position 29809495 bp, A / T mutation at position 30098613 bp, C / T mutation at position 30391267 bp, A / G mutation at position 31955766 bp, G / A mutation at position 32672496 bp, C / T mutation at position 33448952 bp, C / T mutation at position 34565522 bp, G / T mutation at position 34854460 bp, T / G mutation at position 35956877 bp, G / A mutation at position 37299837 bp, T / A mutation at position 37768791 bp, A / G mutation at position 38364999 bp, C / T mutation at position 38772687 bp, A / T mutation at position 39119217 bp, G / A mutation at position 39791727 bp, G / A mutation at position 40956496 bp, A / T mutation at position 42059421 bp, C / T mutation at position 42389769 bp, A / G mutation at position 42831401 bp, T / C mutation at position 43232251 bp, T / A mutation at position 43943244 bp, A / T mutation at position 44224577 bp, A / T mutation at position 44910891 bp, T / G mutation at position 46742409 bp, A / G mutation at position 47231207 bp, A / T mutation at position 47919343 bp, G / A mutation at position 48360872 bp, G / T mutation at position 49359594 bp, G / A mutation at position 49898952 bp, G / A mutation at position 50319548 bp, T / C mutation at position 51049661 bp, A / C mutation at position 51306897 bp, A / G mutation at position 51516277 bp, A / G mutation at position 52191511 bp, G / A mutation at position 52579004 bp, C / G mutation at position 53640899 bp, A / T mutation at position 53976693 bp, T / C mutation at position 54891372 bp, C / T mutation at position 56410343 bp, G / C mutation at position 56902918 bp, A / G mutation at position 57143044 bp, G / A mutation at position 57422435 bp, G / A mutation at position 58005227 bp, T / C mutation at position 58329882 bp, T / C mutation at position 58802812 bp, T / C mutation at position 59036884 bp, G / T mutation at position 60176975 bp, G / A mutation at position 60593132 bp, A / G mutation at position 60929419 bp, A / G mutation at position 61642762 bp,Chromosome 3: T / C mutation at 167511bp, G / T mutation at 444560bp, T / G mutation at 1740957bp, A / C mutation at 2247068bp, A / T mutation at 2663410bp, A / G mutation at 3153958bp, A / G mutation at 3443363bp, T / A mutation at 3918303bp, T / A mutation at 4164736bp, T / G mutation at 4395261bp, G / A mutation at 5114758bp, A / T mutation at 5385938bp, C / T mutation at 5778988bp, G / T mutation at 6150113bp, A / T mutation at 63bp, T / A mutation at 7403440bp, C / G mutation at 8943900bp, T / A mutation at 9256892bp, T / C mutation at 9919466bp, A / C mutation at 10174343bp, A / T mutation at 11633388bp, G / A mutation at 12017984bp, A / G mutation at 12915914bp, C / T mutation at 14184322bp, A / T mutation at 14610187bp, C / T mutation at 15112027bp, A / C mutation at 15742955bp, T / C mutation at 18055450bp, 05bp T / C mutation, A / T mutation at 20806332bp, C / T mutation at 21239659bp, A / T mutation at 21451371bp, A / G mutation at 21908751bp, C / T mutation at 22900014bp, T / G mutation at 23437899bp, C / G mutation at 24068761bp, C / T mutation at 24297401bp, A / G mutation at 25467666bp, A / T mutation at 26854208bp, G / A mutation at 27503027bp, C / T mutation at 27774729bp, T / C mutation at 28167758bp, A / G mutation at 499237bp, G / A mutation at 29844353bp, A / G mutation at 30081511bp, G / T mutation at 30294248bp, G / A mutation at 31271920bp, C / T mutation at 31611643bp, C / T mutation at 31900387bp, A / T mutation at 32420847bp, A / G mutation at 33314059bp, C / G mutation at 33562476bp, A / G mutation at 34537410bp, A / C mutation at 35525002bp, G / A mutation at 36168038bp, G / A mutation at 37291953bp,G / C mutation at position 37938922 bp, T / C mutation at position 38158633 bp, G / T mutation at position 38470830 bp, G / A mutation at position 39158628 bp, G / A mutation at position 39719498 bp, A / C mutation at position 40603131 bp, A / C mutation at position 40865370 bp, T / G mutation at position 41259567 bp, T / C mutation at position 41597965 bp, G / A mutation at position 41910605 bp, T / A mutation at position 42938999 bp, A / T mutation at position 43500081 bp, C / T mutation at position 44213296 bp, A / G mutation at position 44459501 bp, C / A mutation at position 45077111 bp, A / T mutation at position 46562102 bp, T / A mutation at position 47015472 bp, A / C mutation at position 47248636 bp, A / G mutation at position 47755545 bp, G / A mutation at position 48068295 bp, G / A mutation at position 48403665 bp, C / A mutation at position 48859386 bp, A / G mutation at position 49155040 bp, A / T mutation at position 49514429 bp, T / A mutation at position 49801952 bp, G / C mutation at position 50508624 bp, T / C mutation at position 50867456 bp, C / T mutation at position 51172288 bp, G / A mutation at position 51555200 bp, T / A mutation at position 51897141 bp, C / G mutation at position 53046160 bp, T / A mutation at position 53359071 bp, T / A mutation at position 53636779 bp, C / T mutation at position 53869457 bp, T / C mutation at position 54594267 bp, T / C mutation at position 55269130 bp, A / G mutation at position 55572008 bp, A / C mutation at position 55896201 bp, C / T mutation at position 56783882 bp, A / G mutation at position 57104330 bp, C / G mutation at position 57532214 bp, T / C mutation at position 57807917 bp, T / A mutation at position 58451281 bp, C / G mutation at position 58805029 bp, A / C mutation at position 59273801 bp, C / A mutation at position 59815549 bp, A / G mutation at position 60128400 bp, G / A mutation at position 60681360 bp, C / T mutation at position 61275765 bp, C / T mutation at position 61522130 bp, C / A mutation at position 61801828 bp, T / C mutation at position 62069187 bp;, Chromosome 4: C / A mutation at 489947bp, G / C mutation at 886286bp, A / G mutation at 2582998bp, G / A mutation at 3139977bp, G / A mutation at 3595215bp, T / A mutation at 3859888bp, A / G mutation at 4344155bp, C / A mutation at 4566835bp, A / G mutation at 5145569bp, T / C mutation at 6456871bp, G / C mutation at 7321215bp, T / A mutation at 7817588bp, T / G mutation at 8179181bp, G / T mutation at 8910032bp, 6bp G / T mutation, T / G mutation at 9552736bp, G / A mutation at 10022246bp, T / G mutation at 10354612bp, G / A mutation at 10907817bp, C / T mutation at 11734493bp, G / A mutation at 12580554bp, T / C mutation at 13260828bp, T / C mutation at 13765169bp, C / T mutation at 14310601bp, T / C mutation at 16355043bp, G / A mutation at 17630046bp, T / A mutation at 19429489bp, A / T mutation at 19679728bp, and 21079 A / G mutation at 213bp, T / C mutation at 22164490bp, A / C mutation at 22482582bp, T / A mutation at 22780082bp, C / T mutation at 23190102bp, A / G mutation at 23587372bp, A / G mutation at 24228102bp, A / G mutation at 24961164bp, G / T mutation at 25252848bp, A / T mutation at 25844871bp, G / T mutation at 26112318bp, G / A mutation at 27430034bp, C / A mutation at 27889194bp, T / C mutation at 28186129bp, G / T mutation at 862600bp, T / A mutation at 29613029bp, A / T mutation at 29849521bp, C / T mutation at 30074147bp, G / T mutation at 30551177bp, A / C mutation at 31624189bp, T / C mutation at 31868549bp, T / C mutation at 32085476bp, T / C mutation at 33390588bp, T / C mutation at 33885908bp, T / A mutation at 34321828bp, C / A mutation at 34763173bp, A / G mutation at 36691587bp, A / G mutation at 36995284bp,A / C mutation at 37364904bp, C / T mutation at 39303404bp, G / C mutation at 40040754bp, T / C mutation at 40300785bp, T / C mutation at 41007997bp, T / C mutation at 41213469bp, A / G mutation at 41586776bp, A / G mutation at 42194520bp, G / T mutation at 42525010bp, C / T mutation at 42726376bp, T / G mutation at 43723088bp, A / T mutation at 44141447bp, T / C mutation at 44798262bp, A / G mutation at 45230255bp, A / G mutation at 45691471bp, T / A mutation at 45924080bp, A / G mutation at 46335423bp, G / A mutation at 46609807bp, C / T mutation at 47649738bp, C / A mutation at 48054473bp, G / A mutation at 49037444bp, T / C mutation at 49356447bp, T / A mutation at 49823141bp, C / T mutation at 50253140bp, T / A mutation at 50495698bp, G / A mutation at 50782734bp, G / A mutation at 51328829bp, T / C mutation at 51912827bp, T / C mutation at 52524776bp, C / T mutation at 52849059bp, C / T mutation at 53628954bp, C / T mutation at 54121390bp, C / T mutation at 54500655bp, A / T mutation at 54780363bp, C / A mutation at 55043635bp, T / C mutation at 55269248bp, C / T mutation at 56009969bp, T / C mutation at 56360644bp, T / A mutation at 56621645bp, A / G mutation at 57033970 bp, T / A mutation at 57487468 bp, C / T mutation at 58248810 bp, A / G mutation at 58569969 bp, C / A mutation at 58804428 bp, G / A mutation at 59177752 bp, A / G mutation at 59780097 bp, C / A mutation at 60118503 bp, G / T mutation at 60642124 bp, T / A mutation at 60872702 bp, T / G mutation at 61335564 bp, G / A mutation at 61988945 bp, G / T mutation at 62219348 bp; Chromosome 5: C / G mutation at 879645bp, G / A mutation at 1415586bp, C / T mutation at 2111293bp, C / A mutation at 2314335bp, A / T mutation at 2635565bp, G / A mutation at 2920345bp, T / C mutation at 3467779bp, T / A mutation at 3884050bp, C / G mutation at 4393916bp, A / C mutation at 4660080bp, T / C mutation at 5064983bp, G / A mutation at 5762044bp, G / T mutation at 6544275bp, G / T mutation at 6749408bp, G / T mutation at 699 A / G mutation at 7717bp, T / G mutation at 7345402bp, A / G mutation at 7654532bp, A / G mutation at 7932212bp, A / C mutation at 8303258bp, T / C mutation at 8661151bp, T / C mutation at 9176468bp, C / A mutation at 9464239bp, T / C mutation at 9931584bp, A / G mutation at 10539507bp, A / T mutation at 10875599bp, G / A mutation at 11217368bp, C / T mutation at 11702291bp, T / G mutation at 12003330bp, and 12366955 bp A / T mutation, 12808603bp T / C mutation, 13875823bp T / C mutation, 14157737bp A / G mutation, 15150248bp G / A mutation, 15517737bp A / G mutation, 15749958bp A / G mutation, 16257325bp A / G mutation, 16594460bp T / C mutation, 17783174bp T / A mutation, 18355754bp C / A mutation, 18594492bp G / A mutation, 18998718bp C / T mutation, 202 T / C mutation at 53708bp, A / C mutation at 20548723bp, G / A mutation at 22378076bp, A / G mutation at 23213346bp, A / C mutation at 23730799bp, T / C mutation at 23996432bp, C / T mutation at 24241420bp, G / A mutation at 24721774bp, T / C mutation at 26586598bp, A / T mutation at 26893151bp, C / T mutation at 27345167bp, T / C mutation at 27679094bp, C / G mutation at 28185342bp, T / A mutation at 29149844bp,C / A mutation at position 2964933, G / A mutation at position 30587198, C / A mutation at position 30997742, G / T mutation at position 31256011, T / C mutation at position 31705376, T / C mutation at position 32770700, G / C mutation at position 33339763, A / G mutation at position 33573910, C / T mutation at position 34514253, A / G mutation at position 35112239, T / C mutation at position 35669268, T / A mutation at position 36705272, C / T mutation at position 36922640, A / G mutation at position 37131668, C / T mutation at position 37581075, C / T mutation at position 38542724, G / A mutation at position 39044859, G / C mutation at position 39688041, G / A mutation at position 40000656, T / C mutation at position 40395464, G / A mutation at position 41001494, T / C mutation at position 41681731, T / C mutation at position 42174184, A / T mutation at position 42576737, A / G mutation at position 43098562, A / G mutation at position 43487720, A / G mutation at position 43692812, A / C mutation at position 44169923, A / C mutation at position 44373192, C / T mutation at position 44895351, A / G mutation at position 45190288, T / A mutation at position 46681737, T / C mutation at position 47488756, T / C mutation at position 49326052, G / T mutation at position 49639111, C / G mutation at position 50108002, T / A mutation at position 50436257, A / T mutation at position 50696264, T / C mutation at position 50903386, A / G mutation at position 51419745, C / T mutation at position 51669577, T / A mutation at position 52414271, A / C mutation at position 53368505, A / G mutation at position 53605312, T / C mutation at position 53894100, T / A mutation at position 54916254, A / G mutation at position 55477238, A / C mutation at position 55999091, G / C mutation at position 56214132, G / A mutation at position 57070120, T / A mutation at position 57313644, A / C mutation at position 57666549, G / T mutation at position 57877313, C / T mutation at position 58133420, G / A mutation at position 58891833,T / C mutation at 59245611bp, A / T mutation at 59751484bp, A / G mutation at 59976681bp; Chromosome 6: G / A mutation at position 587350, A / G mutation at position 1075497, T / A mutation at position 1679222, G / T mutation at position 2172358, T / C mutation at position 2739350, G / A mutation at position 3206148, C / A mutation at position 3421291, G / A mutation at position 3843416, A / T mutation at position 4064437, G / T mutation at position 4469234, T / C mutation at position 4760643, T / A mutation at position 4992337, A / T mutation at position 5778802, A / C mutation at position 6114864, T / C mutation at position 6400355, G / A mutation at position 6932249, G / A mutation at position 8026496, C / T mutation at position 8885122, T / C mutation at position 9620041, G / T mutation at position 10908040, C / T mutation at position 11117547, A / G mutation at position 12753363, C / T mutation at position 14268050, T / G mutation at position 14586618, G / C mutation at position 14879882, G / A mutation at position 15232542, A / C mutation at position 15807647, C / T mutation at position 16152642, A / G mutation at position 16421643, A / T mutation at position 16738467, C / A mutation at position 17154757, G / A mutation at position 17669050, T / A mutation at position 17995298, A / T mutation at position 18653957, T / C mutation at position 18914180, T / A mutation at position 19545790, C / T mutation at position 20589164, A / G mutation at position 21516044, T / A mutation at position 22028166, A / T mutation at position 22624328, T / A mutation at position 23340429, C / T mutation at position 23832772, A / G mutation at position 24266242, A / C mutation at position 25073811, A / G mutation at position 26222500, A / C mutation at position 26774119, T / C mutation at position 27210621, C / T mutation at position 27689291, G / A mutation at position 28235309, T / C mutation at position 28437764, C / T mutation at position 28668444, T / A mutation at position 29329850, A / G mutation at position 29645718, G / T mutation at position 30524553, G / A mutation at position 31121205, C / T mutation at position 31359984,A / G mutation at 32106045bp, G / A mutation at 32457595bp, T / C mutation at 32753400bp, T / A mutation at 33117124bp, T / C mutation at 33570502bp, T / G mutation at 34176629bp, C / T mutation at 34667060bp, A / G mutation at 38181718bp, T / A mutation at 39546371bp, G / C mutation at 39866670bp, G / T mutation at 40351063bp, A / G mutation at 40681703bp, T / C mutation at 41023800bp, G / T mutation at 41459997bp, T / C mutation at 42016811bp, T / C mutation at 42364688bp, T / A mutation at 42605575bp, C / A mutation at 43099589bp, C / T mutation at 43383766bp, C / T mutation at 43791635bp, A / T mutation at 44792472bp, C / T mutation at 45098188bp, A / G mutation at 45791769bp, A / G mutation at 46506334bp, T / C mutation at 47036051bp, G / C mutation at 47496058bp, C / T mutation at 47812848bp, A / C mutation at 48271374bp, T / A mutation at 49169061bp, A / T mutation at 49429004bp, C / T mutation at 49743523 bp, T / C mutation at 50128359 bp, A / G mutation at 50887722 bp, A / T mutation at 51813382 bp, C / T mutation at 52554583 bp, T / C mutation at 53230845 bp, C / T mutation at 54083112 bp, C / T mutation at 54380581 bp, C / T mutation at 55428291 bp, A / C mutation at 56179611 bp; Chromosome 7: A / G mutation at 603788bp, G / T mutation at 896391bp, T / C mutation at 1282724bp, G / T mutation at 1535834bp, G / A mutation at 1795922bp, T / A mutation at 2124131bp, G / T mutation at 2694802bp, G / A mutation at 3023240bp, T / C mutation at 3227288bp, T / C mutation at 3539581bp, T / C mutation at 3946542bp, T / A mutation at 4409015bp, G / A mutation at 4610418bp, A / G mutation at 5099145bp, 5726 T / C mutation at 774bp, T / C mutation at 6090181bp, T / A mutation at 7174959bp, G / T mutation at 7732660bp, T / A mutation at 7943436bp, C / T mutation at 8179975bp, A / T mutation at 9080480bp, T / A mutation at 9322949bp, A / G mutation at 9569794bp, T / C mutation at 9911214bp, A / C mutation at 10663904bp, T / G mutation at 10877221bp, G / A mutation at 11467271bp, T / A mutation at 11736967bp, and 12053878b p, T / C mutation at 13007410bp, C / T mutation at 13768058bp, A / G mutation at 15198787bp, G / A mutation at 15771972bp, T / G mutation at 16027470bp, G / A mutation at 16465374bp, T / G mutation at 16680508bp, A / G mutation at 17192258bp, T / C mutation at 17558072bp, C / T mutation at 17969940bp, C / T mutation at 18480038bp, C / T mutation at 18732421bp, A / C mutation at 19150831bp, A / T mutation at 8092bp, C / T mutation at 19653197bp, T / C mutation at 20077315bp, T / G mutation at 20839591bp, A / T mutation at 21779850bp, A / G mutation at 23771667bp, C / A mutation at 24586945bp, A / G mutation at 25311728bp, A / C mutation at 25544379bp, T / C mutation at 26417838bp, G / C mutation at 26642847bp, A / G mutation at 26965429bp, A / T mutation at 29758485bp, T / A mutation at 30141591bp,C / T mutation at 30468300bp, T / C mutation at 31400778bp, G / T mutation at 31778180bp, A / G mutation at 32075251bp, C / T mutation at 33230067bp, C / T mutation at 33744108bp, T / A mutation at 36023112bp, T / A mutation at 36396469bp, A / G mutation at 36693769bp, and A / G mutation at 36913195 bp A / T mutation, 37760599bp C / T mutation, 38161672bp T / A mutation, 38702757bp G / T mutation, 39210903bp A / G mutation, 39699534bp A / T mutation, 40385742bp G / C mutation, 41026442bp T / C mutation, 41307086bp C / A mutation, 42969755bp G / A mutation, T / C mutation at 43571847bp, A / G mutation at 43993986bp, T / C mutation at 44394370bp, G / A mutation at 44703340bp, A / G mutation at 45524469bp, C / T mutation at 46045824bp, A / G mutation at 47359820bp, C / A mutation at 47598126bp, A / G mutation at 49292205bp, 50492560 bp T / C mutation, T / C mutation at 50866106bp, C / T mutation at 51554060bp, T / G mutation at 51867069bp, T / C mutation at 52186344bp, A / G mutation at 52386979bp, A / G mutation at 52719201bp, C / T mutation at 53780380bp, G / A mutation at 54228211bp, A / C mutation at 54901003bp; Chromosome 8: G / A mutation at 412562bp, T / A mutation at 985611bp, T / C mutation at 1303653bp, C / T mutation at 1544349bp, T / C mutation at 1922320bp, T / C mutation at 2361255bp, C / A mutation at 2719405bp, A / G mutation at 3000604bp, T / C mutation at 3288220bp, G / C mutation at 3518544bp, T / C mutation at 3933031bp, T / C mutation at 4572621bp, T / C mutation at 5346224bp, A / T mutation at 5602024bp, The mutations at 95bp were T / C, 6172964bp C / T, 6581392bp T / A, 7427921bp T / A, 7653299bp T / C, 10833172bp G / A, 11143937bp C / T, 11563545bp C / T, 12240151bp T / C, 12664163bp A / G, 12976720bp C / A, 13428217bp C / G, 13690299bp T / A, 14184965bp C / A, 145407 A / G mutation at 137bp, A / C mutation at 14927534bp, T / A mutation at 15164050bp, A / C mutation at 16228337bp, C / G mutation at 16839194bp, T / A mutation at 17073823bp, A / G mutation at 17560361bp, C / T mutation at 17805117bp, C / T mutation at 18668691bp, C / T mutation at 19077073bp, A / T mutation at 19321381bp, A / G mutation at 20212862bp, A / G mutation at 20498190bp, T / C mutation at 20748906bp, A / G mutation at 088463bp, A / G mutation at 21591018bp, A / G mutation at 22165379bp, A / T mutation at 23067147bp, T / C mutation at 23844408bp, A / G mutation at 24104507bp, T / A mutation at 24665627bp, G / A mutation at 25079866bp, T / G mutation at 25958988bp, A / G mutation at 27292396bp, G / A mutation at 27511452bp, C / T mutation at 27712853bp, G / A mutation at 27981923bp, G / A mutation at 28485338bp,G / A mutation at 28816819bp, C / A mutation at 29401272bp, C / T mutation at 29733986bp, C / T mutation at 30150021bp, A / T mutation at 30361015bp, A / C mutation at 30665469bp, T / G mutation at 30927726bp, C / G mutation at 31612011bp, and C / T mutation at 31949923 bp C / G mutation, 32350947bp T / C mutation, 32829805bp T / C mutation, 33830538bp T / C mutation, 34428621bp T / C mutation, 34631910bp G / A mutation, 34878333bp C / A mutation, 35167435bp T / C mutation, 35384611bp G / A mutation, C / T mutation at 36295451bp, A / G mutation at 36758686bp, T / C mutation at 38408273bp, T / C mutation at 39084417bp, T / C mutation at 40515577bp, A / G mutation at 40784564bp, C / T mutation at 41770053bp, G / T mutation at 42205041bp, and bp G / A mutation, G / A mutation at 43574397bp, T / G mutation at 44913196bp, C / A mutation at 46413323bp, C / T mutation at 47254739bp, T / C mutation at 47569565bp, C / T mutation at 48443108bp, A / T mutation at 48947202bp, T / C mutation at 50071965bp; Chromosome 9: T / C mutation at 217853bp, A / G mutation at 583042bp, A / C mutation at 796208bp, T / C mutation at 1046351bp, T / G mutation at 1327845bp, G / A mutation at 2838975bp, A / G mutation at 3240786bp, T / C mutation at 3675421bp, G / T mutation at 4651246bp, C / T mutation at 4959856bp, C / T mutation at 5227751bp, C / T mutation at 5933068bp, G / A mutation at 6728623bp, C / T mutation at 7235075bp, 74929 C / G mutation at 85bp, A / T mutation at 7915378bp, A / G mutation at 8155620bp, A / C mutation at 8437140bp, C / A mutation at 9041049bp, T / G mutation at 9320125bp, G / C mutation at 9523788bp, A / G mutation at 10256842bp, A / T mutation at 10516635bp, C / G mutation at 11295005bp, C / G mutation at 11568023bp, T / A mutation at 12159413bp, C / T mutation at 12529731bp, T / G mutation at 12735255bp, and 13042605 bp C / A mutation, A / G mutation at 13493638bp, A / T mutation at 13853217bp, G / A mutation at 14058546bp, A / T mutation at 15219775bp, G / T mutation at 15704220bp, G / A mutation at 16140181bp, T / C mutation at 16412881bp, T / C mutation at 17010061bp, T / C mutation at 17928904bp, C / T mutation at 18193136bp, T / C mutation at 18452448bp, T / C mutation at 18653747bp, A / T mutation at 18878597bp, 196 C / T mutation at 85478bp, G / A mutation at 20221044bp, T / C mutation at 20993779bp, G / T mutation at 21586001bp, A / G mutation at 22002234bp, C / T mutation at 22394787bp, T / A mutation at 22969695bp, A / T mutation at 23876847bp, C / T mutation at 25194999bp, A / G mutation at 25462264bp, A / T mutation at 25741402bp, A / G mutation at 26959004bp, C / A mutation at 27955752bp, A / G mutation at 28258725bp,A / T mutation at 29236123bp, A / G mutation at 29908750bp, C / T mutation at 30127537bp, T / C mutation at 31200772bp, C / T mutation at 31445189bp, T / C mutation at 31759757bp, C / A mutation at 32418276bp, T / C mutation at 33430899bp, G / A mutation at 33857260bp, 34836159 bp A / G mutation, 35910999bp A / G mutation, 36657698bp T / C mutation, 37111454bp A / T mutation, 38333366bp A / T mutation, 38956659bp G / T mutation, 41437696bp T / G mutation, 41655725bp G / T mutation, 42066006bp T / C mutation, 43778385bp G / C mutation, T / C mutation at 44072139bp, C / T mutation at 44341586bp, T / C mutation at 46347906bp, C / A mutation at 46681275bp, A / G mutation at 47089848bp, T / C mutation at 47332573bp, G / A mutation at 47545232bp, T / C mutation at 47787454bp, T / A mutation at 48000048bp, and 48209798 bp T / C mutation, T / C mutation at 49478480bp, G / A mutation at 50204065bp, C / T mutation at 50425019bp, T / G mutation at 52132614bp, T / C mutation at 52785787bp, C / T mutation at 53663195bp, C / T mutation at 54046362bp, T / G mutation at 54605490bp, T / C mutation at 54826243bp; Chromosome 10: T / C mutation at 1534163bp, T / G mutation at 2455069bp, C / T mutation at 3038564bp, T / C mutation at 4197932bp, A / T mutation at 4800809bp, A / T mutation at 5676331bp, T / C mutation at 6642222bp, A / G mutation at 7400941bp, T / C mutation at 7668457bp, T / C mutation at 7897420bp, A / C mutation at 8180708bp, C / T mutation at 8755317bp, G / A mutation at 9215005bp, C / A mutation at 9779335bp, 1040 A / G mutation at 5131bp, A / G mutation at 10801950bp, T / C mutation at 11089097bp, C / A mutation at 11732224bp, T / C mutation at 11959146bp, C / T mutation at 12182036bp, A / G mutation at 12384483bp, T / A mutation at 12977779bp, C / G mutation at 13228019bp, G / A mutation at 13725920bp, A / G mutation at 14297949bp, C / T mutation at 14599462bp, A / C mutation at 15316648bp, C / T mutation at 16226209bp, T / A mutation at 125566bp, G / T mutation at 19323146bp, C / A mutation at 20329041bp, C / A mutation at 20545842bp, A / G mutation at 20831667bp, A / G mutation at 21077840bp, T / C mutation at 21855677bp, C / T mutation at 22076843bp, T / G mutation at 22285822bp, C / A mutation at 22573506bp, T / C mutation at 23139968bp, A / T mutation at 23927295bp, C / T mutation at 24217188bp, T / A mutation at 24958349bp, A / T mutation at 26265143bp, A / C mutation at 26789789bp, A / G mutation at 27013959bp, C / T mutation at 27270677bp, T / A mutation at 27522828bp, A / T mutation at 28230594bp, A / T mutation at 28621223bp, T / C mutation at 28976931bp, G / C mutation at 30161642bp, G / T mutation at 30602940bp, A / G mutation at 30978727bp, T / A mutation at 31649003bp, G / A mutation at 32503776bp, T / C mutation at 33082095bp,G / A mutation at 33398327bp, A / T mutation at 34352656bp, C / T mutation at 34825626bp, A / G mutation at 35322952bp, A / T mutation at 35627155bp, C / T mutation at 35967237bp, C / G mutation at 36252688bp, G / A mutation at 36722144bp, T / C mutation at 37789019bp.

[0026] 2. Identification of parentage in tetraploid long oysters based on the above 1000 SNPs 1. Sequencing of tetraploid long oysters 66 offspring and 2 parents with clear parentage were collected, and 10 offspring and 6 parents with unknown parentage were collected.

[0027] Adductor muscles of the tetraploid long oysters to be tested were collected, and DNA from each sample was purified using the GenoPrep Polysaccharide and Polyphenol DNA Extraction Kit (Shijiazhuang Boredi Biotechnology Co., Ltd.). DNA concentration was determined using a Qubit 2.0 fluorescence quantification instrument, and DNA quality was checked by 1% agarose gel electrophoresis.

[0028] Sequencing libraries were constructed using the GenoBaits® DNA Library Prep Kit for ILM.

[0029] The DNA fragments in the sequencing library were subjected to high-throughput sequencing using an MGISEQ-2000 sequencer (Shenzhen MGI Intelligent Manufacturing Technology Co., Ltd.) in PE150 mode.

[0030] 2. Obtain the genotypes of the above 1000 SNP sites in tetraploid oysters The original data were aligned to the long oyster reference genome, and the aligned data were genotyped using GATK to obtain the genotypes of the above 1000 SNP sites of the tetraploid long oyster to be tested.

[0031] 3. Calculate the IBS and LOD of all parents and offspring to be tested based on the SNP typing results (1) Calculate the consistency of status (IBS) of all parents and offspring to be tested Based on the tetraploid genetic law, the calculation formula of IBS developed by the present invention for parentage identification of tetraploid long oysters is as follows:

[0032] Where n represents the number of successfully typed SNP sites, k represents the kth SNP site, G ik is the genotype of individual i at the kth SNP site, G jkis the genotype of individual j at the kth SNP site, I k (G ik , G jk ) represents the genotype matching score of individual i and individual j at the kth SNP site. If the genotypes of individual i and individual j at the kth SNP site can be matched as parent-offspring, then I k (G ik , G jk ) is 1, otherwise I k (G ik , G jk ) is 0.

[0033] (2) Calculate the likelihood ratio logarithm (LOD) of all tested parents and offspring Based on the tetraploid genetic law, the calculation formula of LOD developed by the present invention for parentage identification of tetraploid long oysters is as follows:

[0034] Among them, n represents the number of successfully typed SNP sites, k represents the kth SNP site, P1 represents the probability that the candidate parent and the simulated parent produce offspring, and P2 represents the background probability (that is, the probability of producing the offspring when the parental genotypes are completely random).

[0035] The calculation formula for P1 is as follows:

[0036] in, represents the probability of the candidate parent gametes appearing, represents the probability of the simulated parent gametes appearing, w represents the frequency of the simulated parent genotype in the population, m represents the number of mismatched genotypes between parents and offspring, which is 0 or 1, and λ represents the mismatch penalty weight, which is 0.1.

[0037] When calculating P2, the genotypes of both simulated parents are based on the population allele frequencies.

[0038] 4. Combine IBS and LOD to identify the parent-offspring relationship When the LOD is greater than 0, the candidate parent is likely to be the true parent, and at least one parent with the highest LOD is most likely to be the true parent of the offspring. At this time, the IBS between the candidate parent and the offspring should be higher than the preset threshold of 0.98.

[0039] When LOD is less than 0, the candidate parent cannot be the true parent.

[0040] In this embodiment, the results of the identification of the paternity and maternity of tetraploid long oysters are shown in Table 1-1 and Table 1-2. The results of the parent-offspring relationship identification between the samples are completely consistent with the pedigree relationship. All offspring with clear parent-offspring relationships have their true parents identified, and none of the 10 offspring with unknown parents have their parents identified. The distribution diagram of LOD between samples with parent-offspring relationships is shown in Figure 1 The LOD and IBS heat maps between some samples are shown in Figure 2 .

[0041] Table 1-1 Identification results of paternity and maternity of tetraploid long oysters

[0042] Table 1-2 Identification results of paternity and maternity of tetraploid long oysters

[0043] It should be noted that the above embodiments are merely examples for the purpose of clearly illustrating the present invention and are not intended to limit the embodiments of the present invention. A person skilled in the art would be able to make other variations or modifications based on the above description. It is not possible to enumerate all embodiments here. Any obvious variations or modifications arising from the technical solution of the present invention remain within the scope of protection of the present invention.

Claims

1. A method for identifying parentage of tetraploid long oysters based on SNP loci, characterized in that: The following steps are involved: (1) Sequencing the adductor muscle of the tetraploid oyster to be tested; (2) Compare the reference genome of Crassostrea gigas to obtain the genotypes of the designated SNP sites of the tetraploid Crassostrea gigas. The GenBank accession number of the reference genome of Crassostrea gigas is GCA_011032805.

1. The genotypes of the designated SNP sites are as follows: Chromosome 1: G / A mutation at 121890bp, A / G mutation at 1057743bp, T / G mutation at 1282310bp, G / T mutation at 1672864bp, T / C mutation at 2377246bp, T / A mutation at 3067043bp, T / C mutation at 3586786bp, A / T mutation at 3999280bp, T / C mutation at 4728772bp, T / C mutation at 5044925bp, T / C mutation at 5300940bp, A / G mutation at 5594188bp, C / T mutation at 5903109bp, T / G mutation at 6330374bp, 6833 T / C mutation at 762bp, A / G mutation at 7943920bp, A / T mutation at 8487382bp, A / G mutation at 9704220bp, T / C mutation at 10224929bp, T / C mutation at 10862004bp, T / G mutation at 11750498bp, T / C mutation at 11990399bp, T / C mutation at 12218512bp, G / A mutation at 12483376bp, G / A mutation at 13361474bp, G / T mutation at 13765448bp, T / A mutation at 14300439bp, T / A mutation at 15205025bp, T / A mutation at 15645 A / G mutation at 080bp, G / T mutation at 15940394bp, T / C mutation at 17045095bp, T / C mutation at 17268995bp, T / A mutation at 17538044bp, A / T mutation at 17899956bp, G / T mutation at 18451368bp, C / T mutation at 18760584bp, C / G mutation at 19501089bp, G / T mutation at 21531956bp, T / A mutation at 22401721bp, T / A mutation at 24526392bp, A / T mutation at 25189059bp, G / A mutation at 25603252bp, A / T mutation at 161161bp, C / T mutation at 26611917bp, A / C mutation at 27004404bp, A / G mutation at 27425048bp, C / T mutation at 27805924bp, A / T mutation at 28632425bp, A / T mutation at 29021720bp, C / T mutation at 33066462bp, A / T mutation at 33718201bp, G / A mutation at 34717118bp, A / C mutation at 35189356bp, C / T mutation at 36608599bp, C / T mutation at 36886136bp, C / A mutation at 37186110bp,G / C mutation at 37483058bp, C / A mutation at 38812679bp, A / G mutation at 39546717bp, C / T mutation at 40317002bp, A / G mutation at 41392051bp, G / A mutation at 43046392bp, C / T mutation at 43389852bp, T / A mutation at 45510791bp, A / G mutation at 46464535bp, A / G mutation at 47037306bp, T / G mutation at 47275653bp, A / T mutation at 47543454bp, A / T mutation at 48596129bp, A / T mutation at 49583230bp / G mutation, T / C mutation at 49868879bp, T / G mutation at 50086266bp, T / G mutation at 50302873bp, A / C mutation at 51214188bp, C / T mutation at 51485365bp, G / A mutation at 52121167bp, A / T mutation at 52424658bp, C / T mutation at 52758029bp, A / G mutation at 53158981bp, C / T mutation at 53542635bp, T / C mutation at 53910234bp, C / T mutation at 54359105bp, T / C mutation at 55808097bp, bp T / A mutation, 57138543bp C / T mutation, 58299726bp G / A mutation, 59081390bp C / T mutation, 59774992bp G / T mutation, 60482812bp T / G mutation, 60889634bp C / T mutation, 61245346bp G / A mutation, 61505117bp C / G mutation, 61817752bp C / A mutation, 62517352bp A / T mutation, 62882914bp T / C mutation, 63105443bp A / G mutation, 64259558bp C / T mutation, 652 C / A mutation at 52225bp, T / A mutation at 66625596bp, A / G mutation at 66835219bp, C / T mutation at 67248469bp, T / A mutation at 67503687bp, A / G mutation at 67906236bp, C / T mutation at 68247170bp, C / T mutation at 68945233bp, T / C mutation at 69565892bp, G / A mutation at 70058662bp, A / G mutation at 70314116bp, A / T mutation at 70765241bp, A / G mutation at 72411555bp, T / C mutation at 73822739bp,A / G mutation at 74137180 bp, A / G mutation at 75053096 bp, A / C mutation at 75443458 bp, A / G mutation at 76271402 bp, T / A mutation at 76958049 bp, C / G mutation at 77190736 bp, T / A mutation at 77491425 bp, A / T mutation at 77966387 bp, C / T mutation at 78744623 bp; Chromosome 2: T / C mutation at 126208bp, G / A mutation at 530724bp, A / T mutation at 1030962bp, C / G mutation at 1883338bp, T / A mutation at 2385548bp, A / G mutation at 2612768bp, T / C mutation at 2905649bp, T / C mutation at 3408267bp, A / G mutation at 4209622bp, T / C mutation at 4735213bp, C / G mutation at 5034762bp, C / A mutation at 5605969bp, A / G mutation at 5935587bp, T / G mutation at 6189866bp, A / T mutation at 484bp, A / G mutation at 7186787bp, A / G mutation at 7919353bp, T / A mutation at 8150271bp, T / C mutation at 8400126bp, A / G mutation at 8695230bp, G / T mutation at 9416767bp, C / T mutation at 9823074bp, T / C mutation at 10397975bp, A / T mutation at 10863126bp, A / G mutation at 11074140bp, A / G mutation at 11296287bp, A / G mutation at 11502471bp, C / G mutation at 12364482bp, and 12574976 bp T / G mutation, T / C mutation at 12794499bp, C / A mutation at 13083777bp, T / C mutation at 13511367bp, T / C mutation at 13877414bp, C / A mutation at 14299527bp, G / A mutation at 14524308bp, A / G mutation at 14736903bp, G / A mutation at 15063383bp, A / G mutation at 15597292bp, T / A mutation at 15867029bp, G / A mutation at 16101395bp, C / A mutation at 16326428bp, T / C mutation at 16905050bp, and 172 A / T mutation at 66489bp, A / T mutation at 17579365bp, G / T mutation at 18124593bp, G / C mutation at 18474645bp, A / G mutation at 18705311bp, C / A mutation at 19267782bp, A / G mutation at 21377238bp, T / A mutation at 22776786bp, A / C mutation at 23709017bp, A / G mutation at 24156723bp, A / C mutation at 24530341bp, G / A mutation at 25094735bp, G / C mutation at 25421687bp, A / G mutation at 25763249bp,C / T mutation at 25980517bp, T / A mutation at 27508872bp, T / C mutation at 28212532bp, C / T mutation at 29113539bp, G / C mutation at 29485500bp, T / C mutation at 29809495bp, A / T mutation at 30098613bp, C / T mutation at 30391267bp, A / G mutation at 31955766bp, G / A mutation at 32672496bp, C / T mutation at 33448952bp, C / T mutation at 34565522bp, G / T mutation at 34854460bp, T at 35956877bp / G mutation, G / A mutation at 37299837bp, T / A mutation at 37768791bp, A / G mutation at 38364999bp, C / T mutation at 38772687bp, A / T mutation at 39119217bp, G / A mutation at 39791727bp, G / A mutation at 40956496bp, A / T mutation at 42059421bp, C / T mutation at 42389769bp, A / G mutation at 42831401bp, T / C mutation at 43232251bp, T / A mutation at 43943244bp, A / T mutation at 44224577bp, 44910891 bp A / T mutation, 46742409bp T / G mutation, 47231207bp A / G mutation, 47919343bp A / T mutation, 48360872bp G / A mutation, 49359594bp G / T mutation, 49898952bp G / A mutation, 50319548bp G / A mutation, 51049661bp T / C mutation, 51306897bp A / C mutation, 51516277bp A / G mutation, 52191511bp A / G mutation, 52579004bp G / A mutation, 53640899bp C / G mutation, 5397 A / T mutation at 6693bp, T / C mutation at 54891372bp, C / T mutation at 56410343bp, G / C mutation at 56902918bp, A / G mutation at 57143044bp, G / A mutation at 57422435bp, G / A mutation at 58005227bp, T / C mutation at 58329882bp, T / C mutation at 58802812bp, T / C mutation at 59036884bp, G / T mutation at 60176975bp, G / A mutation at 60593132bp, A / G mutation at 60929419bp, A / G mutation at 61642762bp; Chromosome 3: T / C mutation at 167511bp, G / T mutation at 444560bp, T / G mutation at 1740957bp, A / C mutation at 2247068bp, A / T mutation at 2663410bp, A / G mutation at 3153958bp, A / G mutation at 3443363bp, T / A mutation at 3918303bp, T / A mutation at 4164736bp, T / G mutation at 4395261bp, G / A mutation at 5114758bp, A / T mutation at 5385938bp, C / T mutation at 5778988bp, G / T mutation at 6150113bp, A / T mutation at 63bp, T / A mutation at 7403440bp, C / G mutation at 8943900bp, T / A mutation at 9256892bp, T / C mutation at 9919466bp, A / C mutation at 10174343bp, A / T mutation at 11633388bp, G / A mutation at 12017984bp, A / G mutation at 12915914bp, C / T mutation at 14184322bp, A / T mutation at 14610187bp, C / T mutation at 15112027bp, A / C mutation at 15742955bp, T / C mutation at 18055450bp, 05bp T / C mutation, A / T mutation at 20806332bp, C / T mutation at 21239659bp, A / T mutation at 21451371bp, A / G mutation at 21908751bp, C / T mutation at 22900014bp, T / G mutation at 23437899bp, C / G mutation at 24068761bp, C / T mutation at 24297401bp, A / G mutation at 25467666bp, A / T mutation at 26854208bp, G / A mutation at 27503027bp, C / T mutation at 27774729bp, T / C mutation at 28167758bp, A / G mutation at 499237bp, G / A mutation at 29844353bp, A / G mutation at 30081511bp, G / T mutation at 30294248bp, G / A mutation at 31271920bp, C / T mutation at 31611643bp, C / T mutation at 31900387bp, A / T mutation at 32420847bp, A / G mutation at 33314059bp, C / G mutation at 33562476bp, A / G mutation at 34537410bp, A / C mutation at 35525002bp, G / A mutation at 36168038bp, G / A mutation at 37291953bp,G / C mutation at 37938922bp, T / C mutation at 38158633bp, G / T mutation at 38470830bp, G / A mutation at 39158628bp, G / A mutation at 39719498bp, A / C mutation at 40603131bp, A / C mutation at 40865370bp, T / G mutation at 41259567bp, T / C mutation at 41597965bp, G / A mutation at 41910605bp, T / A mutation at 42938999bp, A / T mutation at 43500081bp, C / T mutation at 44213296bp, A / G mutation at 44459501bp, C / A mutation at 45077111bp, A / T mutation at 46562102bp, T / A mutation at 47015472bp, A / C mutation at 47248636bp, A / G mutation at 47755545bp, G / A mutation at 48068295bp, G / A mutation at 48403665bp, C / A mutation at 48859386bp, A / G mutation at 49155040bp, A / T mutation at 49514429bp, T / A mutation at 49801952bp, G / C mutation at 50508624bp, T / C mutation at 50867456bp, C / T mutation at 51172288bp, G / A mutation at 51555200bp, T / A mutation at 51897141bp, C / G mutation at 53046160bp, T / A mutation at 53359071bp, T / A mutation at 53636779bp, C / T mutation at 53869457bp, T / C mutation at 54594267bp, T / C mutation at 55269130bp, A / G mutation at 55572008bp, A / C mutation at 55896201bp, C / T mutation at 56783882bp, A / G mutation at 57104330 bp, C / G mutation at 57532214 bp, T / C mutation at 57807917 bp, T / A mutation at 58451281 bp, C / G mutation at 58805029 bp, A / C mutation at 59273801 bp, C / A mutation at 59815549 bp, A / G mutation at 60128400 bp, G / A mutation at 60681360 bp, C / T mutation at 61275765 bp, C / T mutation at 61522130 bp, C / A mutation at 61801828 bp, T / C mutation at 62069187 bp; Chromosome 4: C / A mutation at 489947bp, G / C mutation at 886286bp, A / G mutation at 2582998bp, G / A mutation at 3139977bp, G / A mutation at 3595215bp, T / A mutation at 3859888bp, A / G mutation at 4344155bp, C / A mutation at 4566835bp, A / G mutation at 5145569bp, T / C mutation at 6456871bp, G / C mutation at 7321215bp, T / A mutation at 7817588bp, T / G mutation at 8179181bp, G / T mutation at 8910032bp, 6bp G / T mutation, T / G mutation at 9552736bp, G / A mutation at 10022246bp, T / G mutation at 10354612bp, G / A mutation at 10907817bp, C / T mutation at 11734493bp, G / A mutation at 12580554bp, T / C mutation at 13260828bp, T / C mutation at 13765169bp, C / T mutation at 14310601bp, T / C mutation at 16355043bp, G / A mutation at 17630046bp, T / A mutation at 19429489bp, A / T mutation at 19679728bp, and 21079 A / G mutation at 213bp, T / C mutation at 22164490bp, A / C mutation at 22482582bp, T / A mutation at 22780082bp, C / T mutation at 23190102bp, A / G mutation at 23587372bp, A / G mutation at 24228102bp, A / G mutation at 24961164bp, G / T mutation at 25252848bp, A / T mutation at 25844871bp, G / T mutation at 26112318bp, G / A mutation at 27430034bp, C / A mutation at 27889194bp, T / C mutation at 28186129bp, G / T mutation at 862600bp, T / A mutation at 29613029bp, A / T mutation at 29849521bp, C / T mutation at 30074147bp, G / T mutation at 30551177bp, A / C mutation at 31624189bp, T / C mutation at 31868549bp, T / C mutation at 32085476bp, T / C mutation at 33390588bp, T / C mutation at 33885908bp, T / A mutation at 34321828bp, C / A mutation at 34763173bp, A / G mutation at 36691587bp, A / G mutation at 36995284bp,A / C mutation at 37364904bp, C / T mutation at 39303404bp, G / C mutation at 40040754bp, T / C mutation at 40300785bp, T / C mutation at 41007997bp, T / C mutation at 41213469bp, A / G mutation at 41586776bp, A / G mutation at 42194520bp, G / T mutation at 42525010bp, C / T mutation at 42726376bp, T / G mutation at 43723088bp, A / T mutation at 44141447bp, T / C mutation at 44798262bp, A / G mutation at 45230255bp, A / G mutation at 45691471bp, T / A mutation at 45924080bp, A / G mutation at 46335423bp, G / A mutation at 46609807bp, C / T mutation at 47649738bp, C / A mutation at 48054473bp, G / A mutation at 49037444bp, T / C mutation at 49356447bp, T / A mutation at 49823141bp, C / T mutation at 50253140bp, T / A mutation at 50495698bp, G / A mutation at 50782734bp, G / A mutation at 51328829bp, T / C mutation at 51912827bp, T / C mutation at 52524776bp, C / T mutation at 52849059bp, C / T mutation at 53628954bp, C / T mutation at 54121390bp, C / T mutation at 54500655bp, A / T mutation at 54780363bp, C / A mutation at 55043635bp, T / C mutation at 55269248bp, C / T mutation at 56009969bp, T / C mutation at 56360644bp, T / A mutation at 56621645bp, A / G mutation at 57033970 bp, T / A mutation at 57487468 bp, C / T mutation at 58248810 bp, A / G mutation at 58569969 bp, C / A mutation at 58804428 bp, G / A mutation at 59177752 bp, A / G mutation at 59780097 bp, C / A mutation at 60118503 bp, G / T mutation at 60642124 bp, T / A mutation at 60872702 bp, T / G mutation at 61335564 bp, G / A mutation at 61988945 bp, G / T mutation at 62219348 bp; Chromosome 5: C / G mutation at 879645bp, G / A mutation at 1415586bp, C / T mutation at 2111293bp, C / A mutation at 2314335bp, A / T mutation at 2635565bp, G / A mutation at 2920345bp, T / C mutation at 3467779bp, T / A mutation at 3884050bp, C / G mutation at 4393916bp, A / C mutation at 4660080bp, T / C mutation at 5064983bp, G / A mutation at 5762044bp, G / T mutation at 6544275bp, G / T mutation at 6749408bp, G / T mutation at 699 A / G mutation at 7717bp, T / G mutation at 7345402bp, A / G mutation at 7654532bp, A / G mutation at 7932212bp, A / C mutation at 8303258bp, T / C mutation at 8661151bp, T / C mutation at 9176468bp, C / A mutation at 9464239bp, T / C mutation at 9931584bp, A / G mutation at 10539507bp, A / T mutation at 10875599bp, G / A mutation at 11217368bp, C / T mutation at 11702291bp, T / G mutation at 12003330bp, and 12366955 bp A / T mutation, 12808603bp T / C mutation, 13875823bp T / C mutation, 14157737bp A / G mutation, 15150248bp G / A mutation, 15517737bp A / G mutation, 15749958bp A / G mutation, 16257325bp A / G mutation, 16594460bp T / C mutation, 17783174bp T / A mutation, 18355754bp C / A mutation, 18594492bp G / A mutation, 18998718bp C / T mutation, 202 T / C mutation at 53708bp, A / C mutation at 20548723bp, G / A mutation at 22378076bp, A / G mutation at 23213346bp, A / C mutation at 23730799bp, T / C mutation at 23996432bp, C / T mutation at 24241420bp, G / A mutation at 24721774bp, T / C mutation at 26586598bp, A / T mutation at 26893151bp, C / T mutation at 27345167bp, T / C mutation at 27679094bp, C / G mutation at 28185342bp, T / A mutation at 29149844bp,C / A mutation at 29649330bp, G / A mutation at 30587198bp, C / A mutation at 30997742bp, G / T mutation at 31256011bp, T / C mutation at 31705376bp, T / C mutation at 32770700bp, G / C mutation at 33339763bp, A / G mutation at 33573910bp, C / T mutation at 34514253bp, A / G mutation at 35112239bp, T / C mutation at 35669268bp, T / A mutation at 36705272bp, C / T mutation at 36922640bp, A / C mutation at 37131668bp / G mutation, C / T mutation at 37581075bp, C / T mutation at 38542724bp, G / A mutation at 39044859bp, G / C mutation at 39688041bp, G / A mutation at 40000656bp, T / C mutation at 40395464bp, G / A mutation at 41001494bp, T / C mutation at 41681731bp, T / C mutation at 42174184bp, A / T mutation at 42576737bp, A / G mutation at 43098562bp, A / G mutation at 43487720bp, A / G mutation at 43692812bp, 44169923 bp A / C mutation, 44373192bp A / C mutation, 44895351bp C / T mutation, 45190288bp A / G mutation, 46681737bp T / A mutation, 47488756bp T / C mutation, 49326052bp T / C mutation, 49639111bp G / T mutation, 50108002bp C / G mutation, 50436257bp T / A mutation, 50696264bp A / T mutation, 50903386bp T / C mutation, 51419745bp A / G mutation, 51669577bp C / T mutation, 524 T / A mutation at 14271bp, A / C mutation at 53368505bp, A / G mutation at 53605312bp, T / C mutation at 53894100bp, T / A mutation at 54916254bp, A / G mutation at 55477238bp, A / C mutation at 55999091bp, G / C mutation at 56214132bp, G / A mutation at 57070120bp, T / A mutation at 57313644bp, A / C mutation at 57666549bp, G / T mutation at 57877313bp, C / T mutation at 58133420bp, G / A mutation at 58891833bp,T / C mutation at 59245611bp, A / T mutation at 59751484bp, A / G mutation at 59976681bp; Chromosome 6: G / A mutation at 587350bp, A / G mutation at 1075497bp, T / A mutation at 1679222bp, G / T mutation at 2172358bp, T / C mutation at 2739350bp, G / A mutation at 3206148bp, C / A mutation at 3421291bp, G / A mutation at 3843416bp, A / T mutation at 4064437bp, G / T mutation at 4469234bp, T / C mutation at 4760643bp, T / A mutation at 4992337bp, A / T mutation at 5778802bp, A / C mutation at 6400 T / C mutation at 355bp, G / A mutation at 6932249bp, G / A mutation at 8026496bp, C / T mutation at 8885122bp, T / C mutation at 9620041bp, G / T mutation at 10908040bp, C / T mutation at 11117547bp, A / G mutation at 12753363bp, C / T mutation at 14268050bp, T / G mutation at 14586618bp, G / C mutation at 14879882bp, G / A mutation at 15232542bp, A / C mutation at 15807647bp, C / T mutation at 16152642bp, and 164216 A / G mutation at 43bp, A / T mutation at 16738467bp, C / A mutation at 17154757bp, G / A mutation at 17669050bp, T / A mutation at 17995298bp, A / T mutation at 18653957bp, T / C mutation at 18914180bp, T / A mutation at 19545790bp, C / T mutation at 20589164bp, A / G mutation at 21516044bp, T / A mutation at 22028166bp, A / T mutation at 22624328bp, T / A mutation at 23340429bp, C / T mutation at 23832772bp, A / G mutation at 266242bp, A / C mutation at 25073811bp, A / G mutation at 26222500bp, A / C mutation at 26774119bp, T / C mutation at 27210621bp, C / T mutation at 27689291bp, G / A mutation at 28235309bp, T / C mutation at 28437764bp, C / T mutation at 28668444bp, T / A mutation at 29329850bp, A / G mutation at 29645718bp, G / T mutation at 30524553bp, G / A mutation at 31121205bp, C / T mutation at 31359984bp,A / G mutation at 32106045bp, G / A mutation at 32457595bp, T / C mutation at 32753400bp, T / A mutation at 33117124bp, T / C mutation at 33570502bp, T / G mutation at 34176629bp, C / T mutation at 34667060bp, A / G mutation at 38181718bp, T / A mutation at 39546371bp, G / C mutation at 39866670bp, G / T mutation at 40351063bp, A / G mutation at 40681703bp, T / C mutation at 41023800bp, G / T mutation at 41459997bp, T / C mutation at 42016811bp, T / C mutation at 42364688bp, T / A mutation at 42605575bp, C / A mutation at 43099589bp, C / T mutation at 43383766bp, C / T mutation at 43791635bp, A / T mutation at 44792472bp, C / T mutation at 45098188bp, A / G mutation at 45791769bp, A / G mutation at 46506334bp, T / C mutation at 47036051bp, G / C mutation at 47496058bp, C / T mutation at 47812848bp, A / C mutation at 48271374bp, T / A mutation at 49169061bp, A / T mutation at 49429004bp, C / T mutation at 49743523 bp, T / C mutation at 50128359 bp, A / G mutation at 50887722 bp, A / T mutation at 51813382 bp, C / T mutation at 52554583 bp, T / C mutation at 53230845 bp, C / T mutation at 54083112 bp, C / T mutation at 54380581 bp, C / T mutation at 55428291 bp, A / C mutation at 56179611 bp; Chromosome 7: A / G mutation at 603788bp, G / T mutation at 896391bp, T / C mutation at 1282724bp, G / T mutation at 1535834bp, G / A mutation at 1795922bp, T / A mutation at 2124131bp, G / T mutation at 2694802bp, G / A mutation at 3023240bp, T / C mutation at 3227288bp, T / C mutation at 3539581bp, T / C mutation at 3946542bp, T / A mutation at 4409015bp, G / A mutation at 4610418bp, A / G mutation at 5099145bp, 5726 T / C mutation at 774bp, T / C mutation at 6090181bp, T / A mutation at 7174959bp, G / T mutation at 7732660bp, T / A mutation at 7943436bp, C / T mutation at 8179975bp, A / T mutation at 9080480bp, T / A mutation at 9322949bp, A / G mutation at 9569794bp, T / C mutation at 9911214bp, A / C mutation at 10663904bp, T / G mutation at 10877221bp, G / A mutation at 11467271bp, T / A mutation at 11736967bp, and 12053878b p, T / C mutation at 13007410bp, C / T mutation at 13768058bp, A / G mutation at 15198787bp, G / A mutation at 15771972bp, T / G mutation at 16027470bp, G / A mutation at 16465374bp, T / G mutation at 16680508bp, A / G mutation at 17192258bp, T / C mutation at 17558072bp, C / T mutation at 17969940bp, C / T mutation at 18480038bp, C / T mutation at 18732421bp, A / C mutation at 19150831bp, A / T mutation at 8092bp, C / T mutation at 19653197bp, T / C mutation at 20077315bp, T / G mutation at 20839591bp, A / T mutation at 21779850bp, A / G mutation at 23771667bp, C / A mutation at 24586945bp, A / G mutation at 25311728bp, A / C mutation at 25544379bp, T / C mutation at 26417838bp, G / C mutation at 26642847bp, A / G mutation at 26965429bp, A / T mutation at 29758485bp, T / A mutation at 30141591bp,C / T mutation at 30468300bp, T / C mutation at 31400778bp, G / T mutation at 31778180bp, A / G mutation at 32075251bp, C / T mutation at 33230067bp, C / T mutation at 33744108bp, T / A mutation at 36023112bp, T / A mutation at 36396469bp, A / G mutation at 36693769bp, and A / G mutation at 36913195 bp A / T mutation, 37760599bp C / T mutation, 38161672bp T / A mutation, 38702757bp G / T mutation, 39210903bp A / G mutation, 39699534bp A / T mutation, 40385742bp G / C mutation, 41026442bp T / C mutation, 41307086bp C / A mutation, 42969755bp G / A mutation, T / C mutation at 43571847bp, A / G mutation at 43993986bp, T / C mutation at 44394370bp, G / A mutation at 44703340bp, A / G mutation at 45524469bp, C / T mutation at 46045824bp, A / G mutation at 47359820bp, C / A mutation at 47598126bp, A / G mutation at 49292205bp, 50492560 bp T / C mutation, T / C mutation at 50866106bp, C / T mutation at 51554060bp, T / G mutation at 51867069bp, T / C mutation at 52186344bp, A / G mutation at 52386979bp, A / G mutation at 52719201bp, C / T mutation at 53780380bp, G / A mutation at 54228211bp, A / C mutation at 54901003bp; Chromosome 8: G / A mutation at 412562bp, T / A mutation at 985611bp, T / C mutation at 1303653bp, C / T mutation at 1544349bp, T / C mutation at 1922320bp, T / C mutation at 2361255bp, C / A mutation at 2719405bp, A / G mutation at 3000604bp, T / C mutation at 3288220bp, G / C mutation at 3518544bp, T / C mutation at 3933031bp, T / C mutation at 4572621bp, T / C mutation at 5346224bp, A / T mutation at 5602024bp, The mutations at 95bp were T / C, 6172964bp C / T, 6581392bp T / A, 7427921bp T / A, 7653299bp T / C, 10833172bp G / A, 11143937bp C / T, 11563545bp C / T, 12240151bp T / C, 12664163bp A / G, 12976720bp C / A, 13428217bp C / G, 13690299bp T / A, 14184965bp C / A, 145407 A / G mutation at 137bp, A / C mutation at 14927534bp, T / A mutation at 15164050bp, A / C mutation at 16228337bp, C / G mutation at 16839194bp, T / A mutation at 17073823bp, A / G mutation at 17560361bp, C / T mutation at 17805117bp, C / T mutation at 18668691bp, C / T mutation at 19077073bp, A / T mutation at 19321381bp, A / G mutation at 20212862bp, A / G mutation at 20498190bp, T / C mutation at 20748906bp, A / G mutation at 088463bp, A / G mutation at 21591018bp, A / G mutation at 22165379bp, A / T mutation at 23067147bp, T / C mutation at 23844408bp, A / G mutation at 24104507bp, T / A mutation at 24665627bp, G / A mutation at 25079866bp, T / G mutation at 25958988bp, A / G mutation at 27292396bp, G / A mutation at 27511452bp, C / T mutation at 27712853bp, G / A mutation at 27981923bp, G / A mutation at 28485338bp,G / A mutation at 28816819bp, C / A mutation at 29401272bp, C / T mutation at 29733986bp, C / T mutation at 30150021bp, A / T mutation at 30361015bp, A / C mutation at 30665469bp, T / G mutation at 30927726bp, C / G mutation at 31612011bp, and C / T mutation at 31949923 bp C / G mutation, 32350947bp T / C mutation, 32829805bp T / C mutation, 33830538bp T / C mutation, 34428621bp T / C mutation, 34631910bp G / A mutation, 34878333bp C / A mutation, 35167435bp T / C mutation, 35384611bp G / A mutation, C / T mutation at 36295451bp, A / G mutation at 36758686bp, T / C mutation at 38408273bp, T / C mutation at 39084417bp, T / C mutation at 40515577bp, A / G mutation at 40784564bp, C / T mutation at 41770053bp, G / T mutation at 42205041bp, 42605472 bp G / A mutation, G / A mutation at 43574397bp, T / G mutation at 44913196bp, C / A mutation at 46413323bp, C / T mutation at 47254739bp, T / C mutation at 47569565bp, C / T mutation at 48443108bp, A / T mutation at 48947202bp, T / C mutation at 50071965bp; Chromosome 9: T / C mutation at 217853bp, A / G mutation at 583042bp, A / C mutation at 796208bp, T / C mutation at 1046351bp, T / G mutation at 1327845bp, G / A mutation at 2838975bp, A / G mutation at 3240786bp, T / C mutation at 3675421bp, G / T mutation at 4651246bp, C / T mutation at 4959856bp, C / T mutation at 5227751bp, C / T mutation at 5933068bp, G / A mutation at 6728623bp, C / T mutation at 7235075bp, 74929 C / G mutation at 85bp, A / T mutation at 7915378bp, A / G mutation at 8155620bp, A / C mutation at 8437140bp, C / A mutation at 9041049bp, T / G mutation at 9320125bp, G / C mutation at 9523788bp, A / G mutation at 10256842bp, A / T mutation at 10516635bp, C / G mutation at 11295005bp, C / G mutation at 11568023bp, T / A mutation at 12159413bp, C / T mutation at 12529731bp, T / G mutation at 12735255bp, and 13042605 bp C / A mutation, A / G mutation at 13493638bp, A / T mutation at 13853217bp, G / A mutation at 14058546bp, A / T mutation at 15219775bp, G / T mutation at 15704220bp, G / A mutation at 16140181bp, T / C mutation at 16412881bp, T / C mutation at 17010061bp, T / C mutation at 17928904bp, C / T mutation at 18193136bp, T / C mutation at 18452448bp, T / C mutation at 18653747bp, A / T mutation at 18878597bp, 196 C / T mutation at 85478bp, G / A mutation at 20221044bp, T / C mutation at 20993779bp, G / T mutation at 21586001bp, A / G mutation at 22002234bp, C / T mutation at 22394787bp, T / A mutation at 22969695bp, A / T mutation at 23876847bp, C / T mutation at 25194999bp, A / G mutation at 25462264bp, A / T mutation at 25741402bp, A / G mutation at 26959004bp, C / A mutation at 27955752bp, A / G mutation at 28258725bp,A / T mutation at 29236123bp, A / G mutation at 29908750bp, C / T mutation at 30127537bp, T / C mutation at 31200772bp, C / T mutation at 31445189bp, T / C mutation at 31759757bp, C / A mutation at 32418276bp, T / C mutation at 33430899bp, G / A mutation at 33857260bp, 34836159 bp A / G mutation, 35910999bp A / G mutation, 36657698bp T / C mutation, 37111454bp A / T mutation, 38333366bp A / T mutation, 38956659bp G / T mutation, 41437696bp T / G mutation, 41655725bp G / T mutation, 42066006bp T / C mutation, 43778385bp G / C mutation, T / C mutation at 44072139bp, C / T mutation at 44341586bp, T / C mutation at 46347906bp, C / A mutation at 46681275bp, A / G mutation at 47089848bp, T / C mutation at 47332573bp, G / A mutation at 47545232bp, T / C mutation at 47787454bp, T / A mutation at 48000048bp, and 48209798 bp T / C mutation, T / C mutation at 49478480bp, G / A mutation at 50204065bp, C / T mutation at 50425019bp, T / G mutation at 52132614bp, T / C mutation at 52785787bp, C / T mutation at 53663195bp, C / T mutation at 54046362bp, T / G mutation at 54605490bp, T / C mutation at 54826243bp; Chromosome 10: T / C mutation at 1534163bp, T / G mutation at 2455069bp, C / T mutation at 3038564bp, T / C mutation at 4197932bp, A / T mutation at 4800809bp, A / T mutation at 5676331bp, T / C mutation at 6642222bp, A / G mutation at 7400941bp, T / C mutation at 7668457bp, T / C mutation at 7897420bp, A / C mutation at 8180708bp, C / T mutation at 8755317bp, G / A mutation at 9215005bp, C / A mutation at 9779335bp, 1040 A / G mutation at 5131bp, A / G mutation at 10801950bp, T / C mutation at 11089097bp, C / A mutation at 11732224bp, T / C mutation at 11959146bp, C / T mutation at 12182036bp, A / G mutation at 12384483bp, T / A mutation at 12977779bp, C / G mutation at 13228019bp, G / A mutation at 13725920bp, A / G mutation at 14297949bp, C / T mutation at 14599462bp, A / C mutation at 15316648bp, C / T mutation at 16226209bp, T / A mutation at 125566bp, G / T mutation at 19323146bp, C / A mutation at 20329041bp, C / A mutation at 20545842bp, A / G mutation at 20831667bp, A / G mutation at 21077840bp, T / C mutation at 21855677bp, C / T mutation at 22076843bp, T / G mutation at 22285822bp, C / A mutation at 22573506bp, T / C mutation at 23139968bp, A / T mutation at 23927295bp, C / T mutation at 24217188bp, T / A mutation at 24958349bp, A / T mutation at 26265143bp, A / C mutation at 26789789bp, A / G mutation at 27013959bp, C / T mutation at 27270677bp, T / A mutation at 27522828bp, A / T mutation at 28230594bp, A / T mutation at 28621223bp, T / C mutation at 28976931bp, G / C mutation at 30161642bp, G / T mutation at 30602940bp, A / G mutation at 30978727bp, T / A mutation at 31649003bp, G / A mutation at 32503776bp, T / C mutation at 33082095bp,G / A mutation at 33398327bp, A / T mutation at 34352656bp, C / T mutation at 34825626bp, A / G mutation at 35322952bp, A / T mutation at 35627155bp, C / T mutation at 35967237bp, C / G mutation at 36252688bp, G / A mutation at 36722144bp, T / C mutation at 37789019bp; (3) Calculate the IBS and LOD of all tested parents and offspring based on the SNP typing results. The calculation formula is as follows: ， ； Where n represents the number of successfully typed SNP sites, k represents the kth SNP site, G ik is the genotype of individual i at the kth SNP site, G jk is the genotype of individual j at the kth SNP site, I k (G ik , G jk ) represents the genotype matching score of individual i and individual j at the kth SNP site. When the genotypes of individual i and individual j at the kth SNP site can be matched as parent-offspring, I k (G ik , G jk ) is 1, otherwise I k (G ik , G jk ) is 0, P1 represents the probability that the candidate parent and the simulated parent produce offspring, and P2 represents the background probability; (4) Combine IBS and LOD to identify the parent-offspring relationship.

2. The method according to claim 1, characterized in that In step (3), the probability P1 of the candidate parent and the simulated parent producing offspring is calculated as follows: ； in, represents the probability of the candidate parent gametes appearing, represents the probability of the simulated parent gametes appearing, w represents the frequency of the simulated parent genotype in the population, m represents the number of mismatched genotypes between parents and offspring, and λ represents the mismatch penalty weight, which is 0.

1.

3. The method according to claim 1, characterized in that In step (3), when calculating the background probability P2, the genotypes of the simulated parents are based on the population allele frequencies.

4. The method according to claim 1, wherein In step (4), the method for identifying the parent-offspring relationship of the parent to be tested and the offspring by combining IBS and LOD is as follows: (1) When LOD is greater than 0, the candidate parent is likely to be the true parent, and at least one parent with the highest LOD is most likely to be the true parent of the offspring. At this time, the IBS between the candidate parent and the offspring should be higher than the preset threshold of 0.98; (2) When LOD is less than 0, the candidate parent cannot be the true parent.

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