CN111944913A - Cynoglossus semilaevis disease-resistant breeding gene chip and application thereof - Google Patents
Cynoglossus semilaevis disease-resistant breeding gene chip and application thereof Download PDFInfo
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Abstract
The invention aims to provide a gene chip for breeding of disease-resistant improved varieties of cynoglossus semilaevis, solves the problem that the disease-resistant improved varieties of cynoglossus semilaevis lack of the gene chip, makes up the defects of the traditional breeding technology, and provides a novel molecular breeding method for breeding of disease-resistant high-yield high-quality improved varieties. The gene chip can perform SNP typing of the cynoglossus semilaevis whole genome level, calculate the genetic effect of each SNP locus, estimate the genome breeding value (GEBV) of an individual, and screen fish with strong disease resistance according to the size of the GEBV of the individual; by using the preferred parent fish to breed offspring, the survival rate of the offspring is obviously improved. Therefore, the gene chip of the invention can accelerate the breeding process of the disease-resistant improved cynoglossus semilaevis, shorten the breeding period and provide an efficient molecular breeding technical means for the breeding of the disease-resistant improved cynoglossus semilaevis.
Description
Technical Field
The invention belongs to the technical field of aquatic product genetic breeding, and particularly relates to a gene chip for breeding of disease-resistant improved cynoglossus semilaevis varieties, and a preparation method and application thereof.
Background
The fish culture is a supporting industry in aquaculture, the total output of the aquaculture in China in 2018 is 4991.06 ten thousand tons, the fish culture output is 2693.78 ten thousand tons, and the fish culture output accounts for 53.97% of the total culture output. Meanwhile, fish as a high-quality protein source is also an important component of food composition in China. The cynoglossus semilaevis is a rare and high-quality seawater fish cultured in coastal areas of China, and is popular with consumers due to tender meat and smooth taste. With the gradual expansion of the scale of industrial aquaculture in China, the problems of frequent diseases, degeneration of seeds and the like are increasingly highlighted, and the healthy, vigorous and sustainable development of the aquaculture industry of the cynoglossus semilaevis is seriously hindered and restricted. In order to avoid the overproof medicine residue and the reduction of the quality of aquatic products caused by the abuse of antibiotics in the culture process, the demand of high-quality disease-resistant seedlings is higher and higher. The artificial breeding is a method capable of improving the disease resistance of aquatic animals and simultaneously meets the requirement of developing the green aquaculture industry.
The disease resistance trait is usually a quantitative trait determined by multiple genes. At present, the disease resistance of individuals or groups is generally measured by adopting artificial infection specific pathogens in aquatic product disease resistance breeding, and an optimal linear unbiased estimation (ABLUP) is utilized to calculate a breeding value (EBV) by combining pedigree data. However, the surviving individuals in infection experiments are generally not used as parents for breeding, so that only uninfected individuals can be selected from families with high EBV as parents for breeding. Furthermore, since the disease resistance trait cannot be directly measured, EBV of uninfected individuals cannot be obtained either, which results in a great reduction in selection accuracy and efficiency. Genome selection is a method for estimating breeding value by using high-density SNP markers covering the whole genome, and the method has the advantages of high selection accuracy, shortened generation interval, no need of determining candidate individual phenotypes and the like. Therefore, it is particularly suitable for traits that are difficult to directly measure, such as disease resistance traits. The implementation of genome selection technology relies on high quality SNP markers, and gene chips are a means to rapidly obtain high quality SNPs. The gene chip is prepared by fixing a large number of nucleic acid probes on a support such as a silicon chip or a glass slide, and detecting the nucleic acid sequence of a sample by a hybridization method. The method is already used for colony structure analysis, whole genome association analysis, quantitative trait locus positioning, genome selection and the like, but a Cynoglossus semilaevis (Cynoglossus semilaevis) breeding gene chip has not been reported so far.
Disclosure of Invention
The invention aims to provide a gene chip for breeding of disease-resistant improved varieties of cynoglossus semilaevis, so that the problem of lack of the gene chip in breeding of the improved varieties of cynoglossus semilaevis is solved, the defects of the traditional selective breeding method are overcome, the breeding process and means are optimized, the breeding technical means of disease-resistant high-quality varieties are provided for the breeding industry of cynoglossus semilaevis, the updating and updating of the selective breeding technology of cynoglossus semilaevis are realized, and the efficient and rapid development of the cynoglossus semilaevis is promoted.
The invention firstly provides SNP loci which can cover genome of cynoglossus semilaevis and are related to the disease resistance character of cynoglossus semilaevis and have higher genetic effect value, wherein the SNP loci are SNP loci with sequence SEQ ID NO: 1-SEQ ID NO: 300, at base position 36 of any one of the sequences;
the SNP locus provided by the invention is used for preparing a detection product for breeding of a disease-resistant improved variety of cynoglossus semilaevis;
the detection product is preferably a gene chip;
the invention also provides a screening method of the disease-resistant individuals of the cynoglossus semilaevis, which is to use the gene chip to perform genotyping and screen out the individuals with disease-resistant potential;
the method comprises the following steps:
1) establishing a reference population for disease-resistant breeding of cynoglossus semilaevis, evaluating the accuracy of a genome optimal linear unbiased prediction (GBLUP) estimated breeding value by using the reference population and a chip design site, and comparing the accuracy with the prediction accuracy of an optimal linear unbiased prediction (ABLUP) based on a pedigree;
2) extracting the genome DNA of the candidate parent fish of the cynoglossus semilaevis and carrying out genotyping by using a gene chip;
3) estimating a Genome Estimated Breeding Value (GEBV) of the candidate parent fish by using the constructed cynoglossus semilaevis reference group and the genotyping information of the candidate parent fish;
4) and selecting individuals with disease-resistant potential according to the GEBV of the candidate individuals to cultivate the disease-resistant strains.
The molecular breeding method based on the gene chip can be used for screening disease-resistant individuals of the cynoglossus semilaevis, and the survival rate of the selected progeny of the disease-resistant individuals is improved, so that the breeding process of the disease-resistant improved varieties of the cynoglossus semilaevis can be accelerated, the breeding period can be shortened, and an efficient technical means is provided for the breeding of the disease-resistant improved varieties of the cynoglossus semilaevis.
Drawings
FIG. 1: the distance between two adjacent SNPs of the gene chip of the sole core No. 1 is distributed.
FIG. 2: the gene chip SNP hypo-allelic frequency distribution diagram of the sole core No. 1 of the invention.
FIG. 3: and (3) detecting the genomic DNA of the sample to be detected, wherein M is a molecular weight marker, the rest lanes are detection results, and the sample loading amounts of M and DNA are both 5 mu l.
FIG. 4: and (3) a DNA amplification product detection map of a sample to be detected, wherein M is a molecular weight marker, the rest lanes are detection results, the loading amount of M is 5 mu l, and the loading amount of an amplification product is 1 mu l.
FIG. 5: and (3) scanning results of fluorescence signals after the gene chip is hybridized with a sample to be detected, wherein each point is a hybridization signal.
FIG. 6: and each point of two SNP locus typing cluster maps represents a sample to be detected, and points falling in different color circles represent different typing results.
FIG. 7: and a statistic graph of consistency of gene chips of sole core No. 1 and a re-sequencing typing result.
FIG. 8: genome optimal linear unbiased prediction (GBLUP) and pedigree based optimal linear unbiased prediction (BLUP) accuracy comparison plots.
FIG. 9: average infection survival comparison of families ranked top 5 and bottom 5 for GEBV is shown.
Detailed Description
The invention establishes a method for manufacturing and applying a gene chip for cultivating the disease-resistant improved cynoglossus semilaevis seeds, and aims to provide a novel molecular breeding technical means for cultivating the disease-resistant improved cynoglossus semilaevis seeds.
The following explanations of terms to which the present invention relates are as follows:
SNP: single Nucleotide Polymorphism, abbreviation of Single Nucleotide Polymorphism (SNP), indicates DNA sequence Polymorphism caused by Single Nucleotide variation at the genomic level.
Gene chip: a method for detecting nucleic acid sequence of sample by hybridization features that a great number of nucleic acid probes are immobilized on a silicon chip or glass slide.
Degenerate bases: depending on the degeneracy of the codon, one symbol is often substituted for two or more bases. For example, M represents A/C, R represents A/G, K represents G/T, Y represents C/T, etc.
Reference population: in genome selection, a population with phenotypes and genotypes, the phenotypes being obtained by manual measurement and the genotypes being obtained by genome sequencing, is used to train the model.
Candidate population: in genome selection, a population with only genotype but no phenotype, usually a candidate parent fish in the breeding process, can be evaluated for breeding value by combining a reference population after determining the genotype, and the breeding value is preferably selected according to the level of the breeding value.
BLUP: the Best Linear Unbiased Prediction, abbreviated as Best Linear Unbiased Prediction, is a method for estimating individual breeding values by using pedigree information and a mixed Linear model.
GBLUP: the genome optimal Linear Unbiased Prediction, an abbreviation of Genomic Best Linear Unbiased Prediction, is a method for constructing a genome genetic relationship matrix G matrix by using high-density molecular markers covering a genome so as to achieve the estimation of individual breeding values.
GEBV: genome Estimated Breeding Value, abbreviation of Genomic Estimated Breeding Value, Breeding Value Estimated at the genome level by the genome selection method.
The present invention will be described in detail below with reference to examples and the accompanying drawings.
Example 1: cynoglossus sole core No. 1 gene chip SNP locus screening and chip preparation
1. Establishment of Cynoglossus semilaevis anti-Vibrio harveyi reference population and phenotype data collection
The cynoglossus semilaevis anti-vibriosis harveyi reference groups are all derived from cynoglossus semilaevis families which are bred by a subject group of the inventor for years. Epidemiological investigation shows that vibrio harveyi is a main pathogenic bacterium in cynoglossus semilaevis breeding industry, and infected individuals can have symptoms of rotten skin, rotten tail and the like. In order to improve the resistance of cynoglossus semilaevis to vibrio harveyi disease, the subject group starts to carry out the breeding work of cynoglossus semilaevis to vibrio harveyi disease by a vibrio harveyi artificial infection experiment in 2012.
In 2014, 2016 and 2018, 11,207 tails of 86 families, 3,714 tails of 23 families and 6,141 tails of 80 families were subjected to Vibrio harveyi artificial infection experiments respectively, and fin rays and disease-resistant phenotypes of tested individuals were collected from the experiment. Then, 10-15 samples were selected in equal proportion to the survival rate of each pedigree to form a reference population (table 1), and the genotypes of these individuals were obtained by whole genome re-sequencing.
Table 1: cynoglossus semilaevis anti-vibriosis harveyi reference population
2. Cynoglossus semilaevis reference population whole genome re-sequencing and SNP identification
Extracting and purifying the genome DNA of 1,527 reference populations in the step 1 by using a DNA extraction kit (China, Tiangen), establishing a sequencing library after the DNA quality reaches a sequencing standard, establishing a library type of an Illumina double-ended library (inserting fragments with the length of about 300 bp), and sequencing by using an Illumina HiSeq 2000 sequencing platform.
After sequencing is completed, low-quality reads are filtered, and the filtered reads are aligned to a cynoglossus semilaevis reference genome by using software BWA (http:// bio-bw. sourceform. net /) (NCBI number: AGRG 00000000.1).
Then, the SNP sites were identified using software SAMtools, and 23.57M SNP sites were finally obtained with an average sequencing depth of 6.2X.
3. SNP quality control and gene chip site screening
(1) And (3) performing quality control on the 23.57M SNP sites obtained in the step (2), and removing sites with the Minimum Allele Frequency (MAF) lower than 0.01, the deletion rate higher than 0.1 and seriously deviated from Hardy-Weinberg balance (p ═ 0.001) to finally obtain 2.08M high-quality SNP markers.
(2) The genetic effect of 2.08M high quality SNP markers in step (1) was estimated using the Bayes C method provided by R-BGLR, and the analytical model was as follows:
wherein y is a phenotype vector and 0 and 1 represent individuals who died and survived in the infection experiment, respectively; x is a construction matrix used for connecting phenotype and fixed effect, b is a fixed effect vector and comprises a group mean value, a family establishing place and a family establishing year; miIndicates the genotype of the i-th SNP, aiIndicates the Effect value of the ith SNP, θiIs an indicator variable indicating whether the ith flag has an effect; e is the residual error. SNP effect values were estimated using MCMC-Gibbs sampling, which was performed a total of 30,000 times, with the first 10,000 times discarded as burn-in. After the estimation is finished, the obtained effect values are sorted in a reverse order, and the removal effect value is less than 10-5Finally 751,403 candidate SNP sites are obtained.
(3) And (3) carrying out probe design and evaluation on the candidate SNP obtained in the step (2) by using an Affymetrix genotyping probe design analysis process, and removing the sites with the probe conversion possibility evaluation score of less than 0.6. In addition, under the condition that the SNP coverage genome, the absence of other SNPs in 35bp of the SNP flanking sequence and the GC content of 35bp of the SNP flanking sequence are ensured to be 30-70%, the screened SNP markers are finally used for manufacturing a cynoglossus semilaevis gene chip (figure 1 and figure 2), and the used 300 SNP markers have high genetic effect.
The cynoglossus semilaevis gene chip is manufactured by adopting Affymetrix Axiom chip technology of Thermo Fisher company in America, and each chip can simultaneously detect 24 samples.
Example 2: estimation accuracy of genome selection method using reference population and chip design site
SNPs identical to the chip design sites were extracted from the high-quality SNPs obtained in example 1, a genome genetic relationship matrix G matrix was constructed using these SNPs, and prediction accuracy of the GBLUP method was evaluated using 5-fold cross validation. In addition, the prediction accuracy of the BLUP method is also evaluated simultaneously for comparison with the prediction accuracy of the GBLUP method.
The analysis results showed that the prediction accuracy of GBLUP (0.780) was higher than that of BLUP method (0.599) (table 2, fig. 8). Therefore, SNP loci designed by the gene chip of sole core No. 1 can meet the calculation requirement of genome selection, and can ensure that the genome selective breeding work is smoothly developed.
The specific operation method is as follows (Linux environment):
1. extracting chip design sites from a reference population by using PLINK software; chip site information is stored in a file "cs.
plink--bfile cs.Ref--extract cs.chip.loci.desiged.dat--recode A--out cs.chip
2. The genotype file was generated using R, the code being as follows:
library(data.table)
geno<-fread(“cs.chip.raw”)
geno[,c(1:6):＝NULL]
fwrite(geno,“geno.Rseq.chip.geno”,sep＝“”,row.names＝F,quote＝F)
3. performing 5-fold cross validation by using ASReml-R, a cross validation packet file 'color.random.csv', a phenotype file 'pheno.cs.Ref.dat' of a reference population and a pedigree file 'pedigree.cs.Ref.dat' to respectively estimate the prediction accuracy of the GBLUP and BLUP methods, wherein specific implementation codes are as follows:
the results are collated in Table 2,
table 2: GBLUP and BLUP prediction accuracy comparison
From the 5-fold cross-validation results, the prediction accuracy of GBLUP (0.780) is higher than that of BLUP method (0.599). The prediction accuracy of the GBLUP method is improved by 0.181 compared to BLUP. The result shows that the SNP locus designed by the cynoglossus sole core No. 1 gene chip prepared in the embodiment 1 of the invention can meet the calculation requirement of genome selection and can ensure that the genome selection breeding work is smoothly carried out.
Example 3: method for using gene chip of sole core No. 1 prepared in example 1
Randomly selecting a cynoglossus semilaevis fin ray sample subjected to 24-tailed re-sequencing, and typing the samples according to the following steps:
1. preparation of samples to be tested
Clipping a tail fin of a cynoglossus semilaevis of about 1cm, extracting and purifying the genome DNA of a detection sample by using a DNA extraction kit (Tiangen, Beijing), and detecting the quality and the concentration of the DNA by using 1% agarose gel electrophoresis and an ultraviolet spectrophotometer. After electrophoresis, the DNA is observed under a gel imager, a single band appears, the length of the fragment is more than 10kb, the integrity is good, and the genome DNA quality is considered to be better (figure 3). Furthermore, using a spectrophotometer assay, a detected concentration is considered to be reached if sample A260/280 is between 1.8 and 2.0, A260/230 is greater than 1.5, the concentration is greater than 20 ng/. mu.L, and the total amount is greater than 4. mu.g.
After the genomic DNA of the sample to be detected is prepared, the sample to be detected is processed according to the gene chip detection sample preparation standard flow (https:// www.thermofisher.com /) provided by the U.S. Thermo Fisher company.
Firstly, adding the genomic DNA meeting the detection requirement into a 2mL deep-well plate (96-well plate); adding a denaturant at normal temperature, terminating denaturation after 10min, and opening double-stranded DNA into single-stranded DNA;
secondly, adding an amplification primer, an amplification enzyme and dNTP, sealing a membrane, and then amplifying for 24 hours at 37 ℃; then adding isopropanol with the same volume, placing at-20 ℃ for precipitation for 24h, and fragmenting the amplification product; centrifuging at 4 deg.C and 3200g for 40min, collecting precipitate, and discarding supernatant; then, the mixture is placed in a constant temperature box at 37 ℃ for 20min to remove residual isopropanol; adding the prepared hybridization solution after the DNA sample is resuspended, uniformly mixing, and detecting the quality of the amplification product by using 1.5% agarose gel electrophoresis (figure 4), wherein if the band is single and the brightness is high, the amplification effect is good, and the subsequent hybridization requirement is met.
2. Hybridization of sample to be tested and gene chip
Before hybridization, the hybridization solution mixed with the DNA of the sample to be tested is treated as follows: 10min at 95 ℃ and 3min at 48 ℃; thereafter, the temperature of the hybridization solution was kept at 48 ℃. Taking out the gene chip, immersing the chip in the hybridization solution, and placing the chip in a hybridization furnace at 48 ℃ for hybridization for 24 hours; thereafter, the hybridization results were obtained by eluting, ligating and immobilizing the fluorescent protein, hybridizing the probe and scanning the fluorescent signal (FIG. 5), which was stored in the CEL file.
3. Analysis of typing results
The generated CEL file after scanning was automatically analyzed by Best Practice Workflow of Axiom Analysis Suite (AxaS) software (Thermo Fisher, USA) to generate a VCF file. The analysis result shows that all samples to be tested pass quality control and obtain high-quality typing information (figure 6); 28,016 high-quality SNP loci are obtained for subsequent analysis; among these sites, 63.2% of the sites belong to polymorphic sites, and these sites each contain a SNP site of high genetic effect.
Example 4: typing effect verification of gene chip of sole core No. 1 and application thereof
Typing effect verification of gene chip of sole core No. 1
The typing accuracy of the cynoglossus sole core No. 1 biochip was evaluated by comparing the identity of SNPs obtained from 24 samples in example 2 using the biochip and SNPs obtained from the re-sequencing technique.
The analysis result shows that the average typing consistency rate of SNP obtained by using gene chip of sole core No. 1 and SNP obtained by double sequencing is 94.8%; statistics per site: the completely identical site accounts for 53.5%; sites with consistency of 99-95% account for 15.2%; sites with 95% -90% consistency account for 10.6%; sites with consistency of 90-85% account for 12.1%; sites with less than 85% identity account for 8.5%; sites that were completely inconsistent accounted for 0.2%. (FIG. 7). Therefore, the cynoglossus semilaevis gene chip of sole core No. 1 developed by the invention has good typing effect and typing accuracy, and can be used for accurately typing the designed SNP locus.
The specific method of consistency statistics is as follows (Linux environment):
(1) reading the generated VCF file using PLINK, the commands are as follows:
plink--vcf cs4val.chip.vcf--alleleACGT--recode--out cs4val.chip
(2) map file, named SNPs using "chromosome number: physical location" format, the code is as follows:
library(data.table)
map<-fread(“cs4val.chip.map”,header＝F)
map$V2<-paste(map$V1,map$V4,sep＝“:”)
fwrite(map,“cs4val.chip.map”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
write.table(map$V2,“chip.loci.txt”,sep＝“”,col.names＝F,row.names＝F,quote＝F)
(3) the same SNPs in the re-sequenced individuals were extracted using PLINK, with the following orders:
plink--bfile cs4val.rseq--extract chip.loci.txt--recode--out cs4val.rseq
(4) the consistency statistics were performed using R, and the code was as follows:
application of gene chip of sole core No. 1 in breeding of Cynoglossus semilaevis anti-Vibrio harveyi
Selecting 44-tail cynoglossus semilaevis as candidate parent fish, combining with a reference population containing 1,572-tail cynoglossus semilaevis constructed in a laboratory where the inventor is located, estimating GEBV of the candidate individuals by using a GBLUP method, and then carrying out matching according to a breeding plan. Meanwhile, GEBV of each family is calculated, and the family GEBV is expressed by the mean value of the parents GEBV. Finally, the 44 candidates established 23 offspring families by artificial insemination. When the average body length of the individuals in the offspring families is about 10cm, the Vibrio harveyi artificial infection experiment is carried out to determine the infection survival rate of each offspring family. And then, calculating a correlation coefficient between the family GEBV and the corresponding offspring family infection survival rate to evaluate the feasibility of the application of the cynoglossus semilaevis core No. 1 in the breeding of the cynoglossus semilaevis for resisting vibrio harveyi diseases.
The analysis results showed that the mean values of the pedigree GEBV and infection survival rate of the 5 th ranked pedigree GEBV were 0.15 and 79.1%, respectively, and the mean values of the pedigree GEBV and infection survival rate of the 5 th ranked pedigree GEBV were-0.24 and 58.1%, respectively (fig. 9, table 3). There was a significant correlation between the pedigree GEBV and the survival rate of the corresponding offspring pedigree infection with a Pearson correlation coefficient of 0.71. Therefore, the gene chip of sole core No. 1 designed by the inventor can smoothly implement genome selection and can meet the breeding work of the disease-resistant property of the sole of tongue.
The specific implementation method is as follows (Linux environment):
(1) when the typing results of the candidate individuals are read by using PLINK, the typing results of 2 chips are required to be read because only 24 individuals can be detected simultaneously by 1 chip. The read results are saved in binary form in files xx.bed, xx.bim and xx.fam, with the following commands:
plink--vcf cs.1.vcf--alleleACGT--make-bed--out cs.1
plink--vcf cs.2.vcf--alleleACGT--make-bed--out cs.2
(2) bim file, named SNP using "chromosome number: physical location" mode, code as follows:
library(data.table)
bim.1<-fread(“cs.raw.1.bim”)
bim.2<-fread(“cs.raw.2.bim”)
bim.1$V2<-paste(bim.1$V1,bim.1$V4,sep＝“:”)
bim.2$V2<-paste(bim.2$V1,bim.2$V4,sep＝“:”)
fwrite(bim.1,“cs.raw.1.bim”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
fwrite(bim.2,“cs.raw.2.bim”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
write.table(bim.1$V2,“chip.cs.1.loci.dat”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
write.table(bim.2$V2,“chip.cs.2.loci.dat”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
(3) the same SNPs as the chip in the reference population were extracted using PLINK, with the following commands:
plink--bfile cs.Ref--extract chip.cs.1.loci.dat--make-bed--out cs.Ref.1
plink--bfile cs.Ref--extract chip.cs.2loci.dat--make-bed--out cs.Ref.2
(4) genotype files were generated using PLINK, with the following commands:
plink--bfile cs.1--recode A--out geno.cs.1
plink--bfile cs.2--recode A--out geno.cs.2
plink--bfile cs.Ref.1--recode A--out geno.Ref.cs.1
plink--bfile cs.Ref.2--recode A--out geno.Ref.cs.2
(5) the results of combining the two chips using R are coded as follows:
library(data.table)
geno.cs.1<-fread(“geno.cs.1.raw”)
geno.cs.2<-fread(“geno.cs.2.raw”)
geno.Ref.cs.1<-fread(“geno.Ref.cs.1.raw”)
geno.Ref.cs.2<-fread(“geno.Ref.cs.2.raw”)
geno.cs.1[,c(1:6):＝NULL]
geno.cs.2[,c(1:6):＝NULL]
geno.Ref.cs.1[,c(1:6):＝NULL]
geno.Ref.cs.2[,c(1:6):＝NULL]
geno.cs.1<-as.matrix(geno.cs.1)
geno.cs.2<-as.matrix(geno.cs.2)
geno.Ref.cs.1<-as.matrix(geno.Ref.cs.1)
geno.Ref.cs.2<-as.matrix(geno.Ref.cs.2)
geno.1<-rbind(geno.cs.1,geno.Ref.cs.1)
geno.2<-rbind(geno.cs.2,geno.Ref.cs.2)
write.table(geno.1,“geno4GS.cs.1.geno”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
write.table(geno.2,“geno4GS.cs.2.geno”,sep＝“\t”,col.names＝F,row.names＝F,quote＝F)
(6) using R-ASRreml to estimate the GEBV of the candidate individual, it is necessary to prepare a phenotype file "pheno.cs.dat" and files "names.cs.1. list" and "names.cs.2. list" containing all the numbers of individuals involved in the estimation (reference and candidate individuals) before the estimation, the codes are as follows:
according to the analysis process, the GEBV of the candidate cynoglossus semilaevis individuals is successfully estimated, and after the individuals are bred, the offspring family GEBV and the infection survival rate are shown in the table 3:
table 3: 44-tongue sole candidate individual offspring family GEBV and infection survival rate table
As can be seen from table 4, the average GEBV and infection survival rates for the top 5 family ranked for GEBV were 0.15 and 79.1%, respectively, and the average GEBV and infection survival rates for the bottom 5 family ranked for GEBV were-0.24 and 58.1%, respectively (table 4, fig. 9). The Pearson correlation coefficient between GEBV and infection survival rate of these families was estimated to be 0.71, showing a significant positive correlation. Thus, the survival rate of infection is higher in families with higher GEBV.
The results show that the gene chip constructed by using the screened SNP loci has good typing effect and typing accuracy, and can quickly and accurately detect the SNP loci of the samples. By using SNP obtained by typing of gene chip of sole core No. 1 and combining with GBLUP method, GEBV of candidate individual can be estimated; and then, matching is carried out according to a breeding plan, and the disease resistance of the bred high GEBV offspring seeds is obviously improved. In conclusion, the gene chip of 'sole core No. 1' is a rapid, efficient and reliable genotyping platform, can effectively improve the disease-resistant breeding progress of the cynoglossus semilaevis by combining with a genome selection technology, and can be popularized and applied in the process of breeding of the disease-resistant improved species of the cynoglossus semilaevis.
The SNP related to the disease resistance of the cynoglossus semilaevis on the gene chip of the cynoglossus semilaevis core No. 1 can be used for cultivating the disease-resistant improved varieties of the cynoglossus semilaevis, and has quick, efficient and reliable genotyping effect, so that the SNP related to the disease resistance of the cynoglossus semilaevis on the gene chip of the cynoglossus semilaevis core No. 1 can be used for improving the prediction accuracy of the breeding of the improved varieties of the cynoglossus semilaevis, shortening the generation interval and improving the breeding efficiency by combining with the genome selection technology.
Sequence listing
<110> research institute for aquatic products in yellow sea of China institute for aquatic science
Cynoglossus semilaevis disease-resistant breeding gene chip and application thereof
<141> 2020-08-31
<160> 300
<170> SIPOSequenceListing 1.0
<210> 1
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 1
cctccagatt acaaggttag cattctggta gtgttrgcat aatagctatt tattacagtc 60
aactccctag a 71
<210> 2
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 2
acaatttcgt aactgtaaga acaaatgtcc atgtarttat tgtctgattt tccaagcgcc 60
atagaaacat t 71
<210> 3
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 3
agtctgatct gattatggaa gtaccctgac acgttrgact aatttacgtt gtaggcttgt 60
ctcactgatg c 71
<210> 4
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 4
gttcacacag cagaacaacg tctaaacacc atctgkacca acccagagag ctgaagtggg 60
ctaaatgtga g 71
<210> 5
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 5
aacatgatga aaacaacagc agagacggaa gatgcmagat aaggaggtgt aggaggaaaa 60
gcaggagtaa a 71
<210> 6
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 6
atgagatact gcgtatcagt gatgctagct tcatcragtt tctcttgaag taacgccccc 60
tggtgtttca g 71
<210> 7
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 7
ccctgtattg cattcatgag gaaataacac aatgtygcgt cttatgagga atgttcagac 60
ggttagttta a 71
<210> 8
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 8
gtacagcact agctgtaagt tagctagtgg taaggkgtca ttgagtagat ggtgacaaat 60
cagcataaag t 71
<210> 9
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 9
gacttgatca aacccttgat ccaagacctg ggggcygtga gaacttcaga gcaggtgcag 60
taaagtgtca g 71
<210> 10
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 10
cggttgtagc atcaaggcag cgacgaaagc tgttcrtatc tgtgtgtgtc acctcatgtt 60
gatgtcgtca t 71
<210> 11
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 11
ctgaggattt ttcagtttac taaatgacta actgcracgg tgtatgtttc ggtcccttta 60
atcagacaca c 71
<210> 12
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 12
ttctttcttt cagtgcaaac ttttcattat agcccrtagc tttactgcac cgtgccagaa 60
agtgttatga a 71
<210> 13
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 13
aaacaaatgg tcctgcatcc atcagtgttt ctggcmaaaa aaaaaatcac aaataattca 60
cagtaccaat a 71
<210> 14
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 14
atccagaaga agtgaaacat tttaaactaa gaaagygtag ttgcctctga ttgaactgct 60
tcgaaggaag a 71
<210> 15
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 15
atctggttga tcttcaccag gaccacgttc atgccrtctc tgttcagcag cgtcaggtct 60
ctgtaacact g 71
<210> 16
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 16
cggctttgtc tctcctttcc tctccatcaa tccaaycatc ctgttctttt catctccttt 60
acctcttatt a 71
<210> 17
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 17
ttgtctgtgt ctcagagtgt ctgaatcacg aggccrtaca tgtcgtgtcc cagtcctgct 60
acagcaaaca g 71
<210> 18
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 18
cggtgacagg cctctccgtc gcggggtttg gatcayttcc cccgcgatat tggagtttat 60
ggtttgtaca t 71
<210> 19
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 19
ttagggactt ttgaataatg gtggaaatgt actgamaaaa ctttaaagaa aatagcacta 60
acgaatgtga g 71
<210> 20
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 20
gcagggtgag gagagggcag tacaaagaac tcgtckggcc ctcccactga cccccctgaa 60
ctccttcctg a 71
<210> 21
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 21
tgttaaatta taaaacctaa cacacatact ggtgtkttgt tcagcgtctt tacttgttgc 60
cctatctggc c 71
<210> 22
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 22
tggagtaaga gatagatgtt gtggagaaag atctcyagac agtttagcag acgaaagggg 60
agacatgtga a 71
<210> 23
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 23
ttatcctttt agaagcacat ccagtggata cagacrctga gaaagtgctt tgaaaaggtg 60
cgacagtttt a 71
<210> 24
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 24
acaacagtcg tctgtgctgt gaatggcaat agctgktccc gccccgtccg ttaggccccg 60
atgcctcaca g 71
<210> 25
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 25
tgcaccagct caggtcacac ctggagaaca aacacrtcat gactcagtcg ctgcacagaa 60
tagctcagat a 71
<210> 26
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 26
ggtgccactg gctatttggg acgatatgta gttaaycgcc tgggtgagta ttataatatt 60
tacatgctaa c 71
<210> 27
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 27
gctctcacag tctgacgtgc atgtagcatc cttcarggga tttagtatcg tgggtctgat 60
ctcatcatac g 71
<210> 28
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 28
aggctgtagg tcacagcaga tgaaatgttg ttctcyacag ctgccacttc tcttcttcct 60
gagtaaatga g 71
<210> 29
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 29
gacgtaagca gcacggtcag gaatgacaca cctgckgttt taattgcaaa ctttttcact 60
gtcatggagg c 71
<210> 30
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 30
gtcagtaatt gacgtcgacc gttttctttc cttgartgct aatgaaaatt tgacacgcag 60
cttcttggtg a 71
<210> 31
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 31
actgagccaa actctcaaaa tatagtgaaa ataccygaag ctaggctaag acgtagcatc 60
aaactgtatc a 71
<210> 32
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 32
taaaggaaaa gcggtgcagc aggtaagtgg ctgtgrgcag acacaggact aaatgtcatt 60
caccatcttt g 71
<210> 33
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 33
tcactgtaaa tcagagtcaa acttcaccaa aggggyatta aatgtccaga tacgaacttg 60
tcggtcacat t 71
<210> 34
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 34
agcagacata ccagagtttt cttctccaga cgactrcaat ttagtgtcgg ctccagtggc 60
tttaggtcca c 71
<210> 35
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 35
cagtggccgt ggtctgggga tttgcatttc ttttgyattg cacttttctc cagaaactca 60
ttatagtaag g 71
<210> 36
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 36
atcatcccta accctaaccc ctccctccct gaagcmcttg aatgactgat catgcccaca 60
caagtcatgc a 71
<210> 37
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 37
ccgctctttg gcaacatgtt ccataaacac ctgaaygctc cgaagacaaa agggcttaag 60
cggcgcgctt g 71
<210> 38
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 38
actgatgttt ccgtgtttgg actcatatgt tactgyacgt gtcagacgac taacttattc 60
tgaaaggtca a 71
<210> 39
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 39
gcctctttcc tggaacgaca gatgagtgaa gccccraccg ctgggtttca aatcagagcg 60
cattaaaggc c 71
<210> 40
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 40
cgggacactg ttcgggggtt gtcggaggcg gagtcygtcc agttttagtc agagcagtgc 60
aaggtttacc g 71
<210> 41
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 41
gacgtggaga ggaaggatgg ataaacagag acggayggac agaggagaga gagggtggca 60
gtgatgtgtg a 71
<210> 42
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 42
aaggtgctgc agtgggagaa tctctacgtc gttatragga cagaagcttc gttaacgttg 60
gtttggtgtc a 71
<210> 43
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 43
ccacagactt ctacgcaaat ccattgatgc gatacmaggt cctcaactca gtctgctgtc 60
cagagcagtg a 71
<210> 44
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 44
taattgggaa aagatgggag gaaatcctgc tccccrtcag actctgtctc taggtgcatc 60
tcacaaagac g 71
<210> 45
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 45
gaccctcagc ctgtttgctt tgccatattt tggggmttgg aaagatgcaa gaaaccctta 60
aacctcacca t 71
<210> 46
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 46
cagtgaggct ggttctactc aagagtctgt gacgaygatg atggcacttc caataaaacc 60
cagaagcctt t 71
<210> 47
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 47
tttttgtaaa catcgtgtta tcagactttc ttggcycttc atgtgattta gctcttgcgt 60
atatctcagt a 71
<210> 48
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 48
aaatgcgcat ttgttaacac tggctgtgtt tgtgtkgaat attcctgcaa aaaatatgtc 60
tcacagctgt t 71
<210> 49
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 49
tttgtgcggc cgagccaatt ccatttggct ctgagrgatt tctcaggaag ctcattgtct 60
acgtgacaac a 71
<210> 50
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 50
ctgatggagc taatatgaga cacgtcagca ccatcrccag ggagaagtat ttattggaac 60
ggcgaaagtc a 71
<210> 51
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 51
ggggaggtgg aggagaaggg agttagcgga ggagtrggtc acaccaaagc tcttcactga 60
agcatgaaat g 71
<210> 52
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 52
aagaacagac acatgttgag agagaggtca ttgccrctct gtgattgttg ctaacataag 60
ttctgctcat g 71
<210> 53
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 53
ctaagtaaaa cacaacaagt tagcataact aagacycact ttgtcagtca gtagtttatg 60
tcaggttggt g 71
<210> 54
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 54
tctcaaacac tccagaataa agagatggaa acacamtcac cctttggatt aaacctctcc 60
acacagatgc t 71
<210> 55
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 55
gtttctttga tcaaggatct ttcatggaga ttatgmagcc gtgggcccag agtgtggtgg 60
taggcagagc c 71
<210> 56
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 56
ctcgccggct aatacatata ttgattgttt gctggrgaga gaagcagttt ttgaagctgt 60
ggaagagatg g 71
<210> 57
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 57
actgtaaact gcaagactgt aggcatggtt tacacrgaca catatatatg cctgacatac 60
tgacccaaat t 71
<210> 58
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 58
ttgcagtttt tgcagaatga agccattatc taatckaact aaattttgtt tttgtttttg 60
tgaaagatgc t 71
<210> 59
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 59
atgagaaata agtgtgagca gatccacagt agaacrcccg ttgctgccga tgtctcaaca 60
ttcatacaag a 71
<210> 60
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 60
tgtaaagtct agagcttgat ggtttcccag acttgrgatt gggaatcgca ttttttgaac 60
cttttataac t 71
<210> 61
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 61
atttcatata tcactaatgc ttgggcaaga aagtcrttaa atccgacgcg aatgctactc 60
atggtggcag t 71
<210> 62
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 62
tggatggatt gacctggata ggccgctatt tccccrgttt tatttcactc cctatgcttc 60
tctattagtc t 71
<210> 63
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 63
actgggtgaa gcacttgact gatgtttctg cgattyggca ataattaaaa gcatcagtga 60
ccaacaggaa a 71
<210> 64
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 64
ttgatgagca agcacttgaa gctctcaagc acgaakgaat ggtacagcca gctaaagtac 60
agttggactt t 71
<210> 65
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 65
cttctccttt aggttcaata gaccttcaga ggagcmgagg tgaagaaacg ccaacatttt 60
gatcacagcc a 71
<210> 66
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 66
ttcctgggct gataatagcc agaagatcag ttaccrtgtt ttacagactg tgttgagtaa 60
tggcagtggc a 71
<210> 67
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 67
taagacttgt tgttcagaca ggaagcataa gttgarctgt gggagaaaac gctgcagccc 60
gtgaatcata t 71
<210> 68
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 68
ctatatcaga aactagacaa tcagaagctg gactayggga tgaattatga gcaaagaaat 60
gttgaaaaaa g 71
<210> 69
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 69
agctcctgac gttggtgatc ggtgtacacc actctrtact tgtccttcgt gcgagtcttt 60
ccaactgcag c 71
<210> 70
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 70
tgtcgactgt actcaacctg acacacacaa acagaygatg gacaccctgg tcactcaaca 60
tcttcaacac t 71
<210> 71
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 71
ccgcagaaaa ccatttttcc ggctgtaact tcaacycctc caaacttcgc agagcattat 60
ctcattagtg t 71
<210> 72
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 72
ctgcacttgc tacggaggca aacatgtagg atgtcrtccc ccgtcttttg tttagatgca 60
atgatgtcct t 71
<210> 73
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 73
ggatggaaag aatgtcagtc gggactggtg acagtmgacc tggtgatatg tcactgttgg 60
atcattttaa t 71
<210> 74
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 74
cacgccatca accctatgat gtcacacagt tcaccrcatc atcctcagaa cagcgcttca 60
ccacccagta a 71
<210> 75
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 75
taggctgtta aacttgtcag atgatatcag gttaarattt gtatttatta tgtactgcat 60
caggttaggg g 71
<210> 76
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 76
ttcagacagc tcacatttcc atactgctac ttatgycgga taactcgcat acatataata 60
catcattgag c 71
<210> 77
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 77
ctctgtccca ggatttatac gtggcagatt aacagrgtga acaccgtaaa agagacaaat 60
ttcaaccaca a 71
<210> 78
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 78
cagccgtcaa tgacggccat tgttcatgac acgacrcaaa cgtatgagcg ctgacaagtg 60
gatcagctct c 71
<210> 79
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 79
gtggacagca gggttagaca aatgcaacac acatgyatac acacacacac ttacatacac 60
acttatacac a 71
<210> 80
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 80
cgcctgtggt gcagaagtgt ttcctgtttg ggtccmagac acattcgctg aggtcacaca 60
gtcacttctt t 71
<210> 81
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 81
tctctagttt atgcttcaca aaaggtttat tgagtrccaa agattttcca cctacattga 60
cattcgtagg t 71
<210> 82
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 82
cgatgccact ctccgacccc actgttccat agtgcrctgt gctgaaatcc aaactttgct 60
agaatggtga a 71
<210> 83
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 83
tgtctctctg tgtcttctcc cctcctcccc tcctcmccct tttactgtta tctcctattc 60
ttcctcggac t 71
<210> 84
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 84
atggggagtt caaaagagac ctgcgtcgcc aaattrcagc tttgactaaa ggccccttcc 60
tctggtttaa a 71
<210> 85
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 85
caggtgcagg ggttgacaca gctggttttg atgtargttg tgttgttcaa agatataatt 60
catgctgagg a 71
<210> 86
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 86
atgaagaagt aagcacagaa ttcctcttta atgatyaaat tcatacaaat ttaacatttt 60
taaattcaaa a 71
<210> 87
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 87
cagacggctc gcgtcccaga aaaagaacgt ggtacygagc caagtgcatg tgtacgccaa 60
accgattcat a 71
<210> 88
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 88
ccaacacaca caaaaaacta aacaaaaaca caagckcgtg agtgttgatc aatagcattg 60
gtttgtgacg g 71
<210> 89
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 89
cttgatgcta cgcaatgttg tggccagaaa cttcarctct ttttcctgtg tgtagctgta 60
agctaggtca a 71
<210> 90
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 90
gtcagcatac attctggctt catgatgtgc tctgtyggac gtctcaagtc ccagcttctg 60
catgtaatgg g 71
<210> 91
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 91
cacaatttac acctctccaa atgtcaaatg tctgaygctc ttaaatcctc tttgattgta 60
tctttgcagt t 71
<210> 92
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 92
atagattctt ctcttagggc agcacgtgcg taccartgag attgcaattt ccggctgaat 60
aatgacaacg g 71
<210> 93
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 93
aaatgagtca cagacatggg tcacagacat gggtcrcaga catggactga ctggcgctaa 60
ctttgaccgc t 71
<210> 94
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 94
catgcagcta attatttcaa aagtggaaag aaaatkaatc ttttcaatca gaattctctt 60
acaaataata t 71
<210> 95
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 95
gaaaacgtta gagacggatg gtcaattaaa aaggargaga agaagaaacg aaggaaggac 60
gtccacctcg g 71
<210> 96
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 96
cttctgccat aaaaacaaca gaaacctcat caaacyggag ccgcgtggaa tcaaacgctg 60
ccgtcaatga a 71
<210> 97
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 97
ttaaggcaac ccacctactc cgtggccgta ctctgyggtc tgagaaagct tgtttttaaa 60
ccgtttaaac t 71
<210> 98
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 98
ctaccatgaa cattatcagg ctcatccggc agaccrggac acaacagcag catcacatgg 60
agcttctgca g 71
<210> 99
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 99
tccgggcgga gagagaaagg tgttgtgtaa gaaaamacaa acagctctgc agacagactg 60
agtgatgagt c 71
<210> 100
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 100
catctccggc aatgctgcaa agaaagactt gtagcrctgc aggtacatca aaaactccct 60
aaaataaaat a 71
<210> 101
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 101
gtgattctct tatctttgag gagtaaaaca ttttgyactc tggcctttaa atgacctttt 60
gttcgacaca a 71
<210> 102
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 102
ataaatgatt agcctgtgaa tgtgaggtaa cacagyggag tgaatcagat ctaacattat 60
catctgataa a 71
<210> 103
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 103
tcaacgattt aattccagga atgatgttgt caaacygtcg ctctgaaact gacttgcgta 60
accaaacaat t 71
<210> 104
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 104
acgactgtac gtttatttag gatttaccca aaagtmattt ttaggagaat tctagctgtg 60
taagctggaa a 71
<210> 105
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 105
gttgcctttg ccttctgaga aacatctccc ttctgyagca gagtcctttt cagagcctgt 60
cagatattcc a 71
<210> 106
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 106
ggtgtccttc tgtgggacaa cagagaagat tccacrcagt ggagttagca cagactagtg 60
actgccagtc a 71
<210> 107
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 107
acagcagaag tcttatttca gccattaaaa aatccygcac tttgaaattc cctgccaaag 60
acaaccaggt a 71
<210> 108
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 108
gaggtggttc tcctcagggt caggttaata ctgccrtctc catttccatt gtgggtggac 60
aaggagccgg a 71
<210> 109
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 109
tactctgccc ccgtgtggac aaccggtgta taaagygtgt ttataatatg ctaatgttga 60
agatgacgta g 71
<210> 110
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 110
caatattgat gcgataacct gaccagccct aaaatktaat acttattaaa aataacaata 60
tttattaaaa a 71
<210> 111
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 111
ggctggacac ggtgatgcaa caagaagaac agcaamcaaa aagaaagatt cttaatgaga 60
ccaggaagac t 71
<210> 112
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 112
cttcattagc gaggaattct cttgactcct tgaacygcat acaatatttt gaacaatctc 60
tttgcattct t 71
<210> 113
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 113
gacttggaag aagaccatag tgtaacctct ccaacrccaa acttttatca tttcctaagc 60
cagaaagtgt c 71
<210> 114
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 114
gagaagtgaa ccacgagaga aggtccttca aggaamaagg aacgtacacg aagaaagggg 60
ctgtctgtgt g 71
<210> 115
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 115
agcctgtgcg gcacatttct tcctttgaag taccgyattc atgtctgttc ttttctgttc 60
gcgctacgtg c 71
<210> 116
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 116
gttttaatgt cccgtgttta gatggctcca ccttcrtctt ccatggcaac gtgattcaga 60
tttcacacag c 71
<210> 117
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 117
ttaagttaat acgattgaaa tcaggtagta aacctyggtc tcacaactca gggccattat 60
tattagctat a 71
<210> 118
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 118
cggtaatggg cttgacagat aaccttgagg tctgcygtgc tgccgtctca gggcaacacg 60
ccactgctcc t 71
<210> 119
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 119
ccttttcatt ctgtaagtgt ttctattgac attttkggcc tatttttgct gtgttgtagt 60
tcaaaagctt a 71
<210> 120
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 120
cagctggatc ttcacagcta tttgatctgt cgcagmgcaa tctgagaagt gtacagagcc 60
atgaccttct c 71
<210> 121
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 121
cagacctaca tacatgtcgc gtatacaacg tctacycgcc cacaaaacag aaatatagaa 60
ataatttggc t 71
<210> 122
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 122
ttcatccaac tataatcatg gaaaacacat cttgtrtggt tgcgccctgt ttccatcatt 60
ccatgagtcc a 71
<210> 123
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 123
gaagtcccga gttcaggtat gcctttaagg agatcytgtg tttgaagagg aaccctgggc 60
cggagaacgg a 71
<210> 124
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 124
cgacaacaaa tctgacttga gtgataaggt tttctkggtc tcccggggag ggttgcattg 60
actcattaga c 71
<210> 125
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 125
tgaagagtaa tgatgaaggt gggtggacaa agggamgaga gacagagtga gacatgccac 60
acgtcatgtc c 71
<210> 126
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 126
gagggagcag gaggacgggg aacatgtttc cagccrctgt ggccttgact tgttgttaac 60
tcaacagtag t 71
<210> 127
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 127
ttgccagatg ttacaaatct aactgtgcat tgatcrgtga cgtgatagcc catagagatt 60
gcttcatttg c 71
<210> 128
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 128
tgtggctgca gataatggat ggctttcata aatggygtgg agctttgcac gggggaggca 60
gaaatgcagc g 71
<210> 129
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 129
ttcctctgtg atgccaacct gtttctgctg acagtyccca cacacaacag gcaacaggca 60
gtgggcgaga g 71
<210> 130
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 130
gtagtagcga gccagaccag ttgcactacc aaaaayctgc cacagaaggg ttgggtcttc 60
ttctctgttc t 71
<210> 131
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 131
ctgagaagca gaaacagaga cagtgagaga cctggrgaag ccgaggggag agaggaggcg 60
gaggaggctg a 71
<210> 132
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 132
agtgaaaaag caatttgatc catgttacaa cagttyattc agggttagag taatttattg 60
caccctggct a 71
<210> 133
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 133
agaagattcg acaactcttc agagaaacca agctargacg cagacttggc tgacaactaa 60
ctactaacta a 71
<210> 134
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 134
cgccgcctgt tcaagagaat aaaaaaattg tctgargcga aaatcacgtg ttagtggtac 60
agtagtgcgt t 71
<210> 135
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 135
ttgcttgttt tacacatcag gactcatcag attagygact ttgctgcaga atgatgtgtt 60
tgtgtgaatg c 71
<210> 136
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 136
tttcacaaat gaatgtgcgc ataacggtaa taatgkcatc gatgatgtcg gactaacagc 60
ggtgagggtt a 71
<210> 137
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 137
caacatcata aatcatccgc taaatcttta atctgmgaaa aaaaaaacct ctcacccagt 60
gaggacctgc g 71
<210> 138
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 138
gacgctgatt gttctcacac tcctttctcc tccttytttc agtctgatcc tgttgacttt 60
ttctactcca g 71
<210> 139
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 139
ctcctcctaa tgagcagttt attcaaaaag caattmattg accacaagag gtgacggcag 60
ctcggcttta a 71
<210> 140
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 140
actgctgcca caggacgaag gtcaggatta cagggyccta tttgttccca ttgtcaccct 60
gacgacctga c 71
<210> 141
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 141
aagtttaaat gtactgaaaa aagatttggt gttttrcagc caaacttact gcgtttcttc 60
ttgttcattg t 71
<210> 142
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 142
tagaaatatc ctgatcaggc cctttatgct tcctaygttt tacagagtta aaaaggaaaa 60
acaaagaatg a 71
<210> 143
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 143
ttcattattt aatattgagt aattgaaatc cagacycacc agacgcctga ctccatgttg 60
tatgcccaga t 71
<210> 144
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 144
ctgtgtgggt gcgagcgtgt ttctgctttt gtccckcgtg tgtgaatgcc tccacctcta 60
gtgtctctag t 71
<210> 145
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 145
ggtgaatgct gacttcaggg ttacgtgttc ttgatkgctt cattttgtgc cgtttgcaaa 60
ttctaagtgt c 71
<210> 146
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 146
taattagccg tgcctctcca tgtgaaccga accagrgctg tgtttcagtt tgttgcctcg 60
cctttctcca t 71
<210> 147
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 147
tcagtcttct tcaaatagcc aggtttgttg tctcgyagta aaagccaaca gtgagaatca 60
ggtccacaca g 71
<210> 148
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 148
gttgcacatc ggagagtttg acacctttta aaaacycagt tattatgtca aggctaagtt 60
ttcggtgaca t 71
<210> 149
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 149
gtaacagcag ctggtctaat tctgacagac atcacytgaa gctttgcctt aacacagatc 60
tgaaatccac c 71
<210> 150
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 150
atttttccct gaatcaacag aaacaactgt ttgatrttca acattgtgcc ctaaggcttc 60
tgtacatagt g 71
<210> 151
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 151
ctcacagtga aactgtacgg cattgaggcg ttctgygagg aacacacaga gagagaggga 60
gagagagaga g 71
<210> 152
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 152
caggatgact ccgtcgcaaa ctgtcaggaa ctacartcac gagcagagca gccaacgtca 60
gagtcgggag t 71
<210> 153
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 153
ggatggccat ggcgataaga gcgatgtagg aaaacrgggg tttgggtctc cgtgtgtagg 60
gcttggactt g 71
<210> 154
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 154
tttacaatgt ataattattc aaaacataca aagccrgagg ttgatattac atatggcaat 60
tatttgttta a 71
<210> 155
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 155
ggcaaatcac gtatgactct tttgaagtgt agcagmaaaa aaaaaaaaaa aagtaaaaca 60
ctgaccatgt c 71
<210> 156
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 156
ggttcttcag aaccttcagc agctgaggcc tcctcygact tcttctcttc ctctaactcc 60
tcaggcgagg t 71
<210> 157
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 157
acgctgcatg tgggacggaa tgtgttcaca gattgrcagc ctggacttct cctctataaa 60
tggataccac g 71
<210> 158
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 158
tgctgaaata gtgtagtcgg actgttgttt atcccygttc atcgcctctc aggttaccca 60
gctgctatgt a 71
<210> 159
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 159
gcccttcgat tatctgatac agctcctgag ccttgrcgca tgaatcacag gaagtagaat 60
tatttccaga c 71
<210> 160
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 160
aaaaagagga atagatgtgg aagaattaaa tagttyattt gtggactaat atcctcagta 60
acactggaaa a 71
<210> 161
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 161
ctttgatcgg atgtttcccc gagtcgaact ctcacmgttg gtccgttgtc aggatcattt 60
ggtggcgatt g 71
<210> 162
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 162
gaatggagct ggaggccaca aaatctgtac gtacgrttta tttgattatt tattggattt 60
ttccacactg t 71
<210> 163
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 163
cattattatt ctgcccatct gtgcattata tgtcayctgg agctttaatg tagaacctaa 60
tctaatctaa a 71
<210> 164
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 164
cgaccctggt gttgacctgg tctcaggacg tccacyttgt ttcttaggaa tgagctttaa 60
atgtcggatg a 71
<210> 165
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 165
gagccctgtg aagaagacat tggaagtttt gaaagygccc atatagcaat atgggggctg 60
aaatgtggca t 71
<210> 166
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 166
gatatttcct ctgctgttca acatcaccgt tccacraaac gaacacataa caatagccga 60
actgcatctt t 71
<210> 167
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 167
accataccat gcaaactatt cattagcata gccacraagc tacattagca caactacgtc 60
aatataatca c 71
<210> 168
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 168
ggtgcagata attaacatca gggctgtgag aaaatrtcga tacgacgata tatcgcgata 60
tttgtttttt g 71
<210> 169
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 169
acgcacgaaa aaaagatgag tgtatccaac tttccycgac gagggtgaag tgtctgcgcc 60
gaccacagag c 71
<210> 170
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 170
ggggattttc aggtaattca aggacacttt tccccragca cttgcttttt ttggaccatt 60
ttgaggatac a 71
<210> 171
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 171
gctgtgatcc gtctgggctt gatggaaatg atagcmtttg ccaaactctt ctccaaggac 60
aactctgttc a 71
<210> 172
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 172
tctgcaaatc atcaattttc tctgagcctg ggaagmcttg ataacatcgt tctcacttgc 60
agcatctgtg a 71
<210> 173
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 173
ctgctggagc tctgattatc ctcctcacac ttttgktctt caacttgaca ccattgaccc 60
cttctaatat g 71
<210> 174
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 174
tccgagagtc accggagacg gctggacaag gactcrggat ttggttacag ttggaacaga 60
agagacagca g 71
<210> 175
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 175
gtaacaggtt ctgcatgaga ccctatcaca ggaaaytgca aggattaaaa gagagcctgt 60
gggaagaagt a 71
<210> 176
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 176
cctcaaaaat tgggccgtag tggacgaacg gccccrggtt gcgttgtttg ttttactgtc 60
acaccgggcc a 71
<210> 177
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 177
ccagatacag agctgcagcc tgggctggac aatagragct caaaggtcat cagcctgcca 60
gcagatcggc c 71
<210> 178
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 178
tttggggaag gcagcttaga aaatagatac atttcmtgtg ttaatttatg ttctgaacgg 60
ttatgacctt t 71
<210> 179
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 179
cagatgtcaa ctacaccggg atgttagcta gcaacrtgag gactcgtcgc atattaacct 60
tctctgagag c 71
<210> 180
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 180
tggcggcata ttgtaagcga cgtcttgtcg atgcgyggct ttgtccacgc tgtcagtttt 60
acttcatcaa a 71
<210> 181
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 181
gctggatcca ggcccagcag aagatattga agatgktctc agccactggt cctgtccttc 60
cacgatgagt g 71
<210> 182
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 182
tgcaagatga agcagctctt ccatacaagg tcacartgtg attcattgtc aaccaaacct 60
attgtcaacg a 71
<210> 183
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 183
ctgcgaagtg ctttatgaca ggttcactga ggaagraaga agaaaagctt catttaaacc 60
cttaaataca g 71
<210> 184
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 184
taagatacac tatctgttaa gttttttaag tattgrcttt taaacccttt cttccttcaa 60
ataaaactag t 71
<210> 185
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 185
ggaggaggat ggaggagaaa aacagagtaa agtagrcact gacaagacag aggagggctg 60
tggtaagttt c 71
<210> 186
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 186
tactgtatgt atgcttaata catgggcaaa aaagtyccca aaagcagaac acaaatgaaa 60
gaaagtgaat g 71
<210> 187
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 187
taatggcatt tgtttccagg cagtgtcctt taccamcgac ccatgacatt ttttgtgaag 60
gtctgagtca c 71
<210> 188
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 188
gcagctgaag ggagaggtca catctgcatc tgtgcrtatg agtctttggg agttggagga 60
tcgttgttgg a 71
<210> 189
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 189
catcggtgtt ttctatggtt gtatgttgga caacaycgtg aaaaccttta tgcagatttt 60
acctcttgta g 71
<210> 190
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 190
tcagacttca aaagtgaaat tagtcgtgtg tgttgmgcgt cgcagcgagg cggcagcgtg 60
agggaattaa a 71
<210> 191
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 191
ccactgtgct aaaatctgtg acctgatact tactamaggg ttgagtttta aattactctg 60
gggcttgtta t 71
<210> 192
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 192
gccggtgtca tctgtaacat ctcgcattgt tggccktgca gagtaaatgg gagcatgtat 60
aggggtattt g 71
<210> 193
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 193
accatctgtg tatgaagtga aacatctctg agtgcycaga ccagagcagc tgacagtaga 60
ggggctgcag c 71
<210> 194
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 194
tgtgatgatg atgatgatgt gtgaattctg agtccmtctg ttgtattgta ttaagcacat 60
ctctgtacct c 71
<210> 195
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 195
ttccagagat gtcagcctgg aatttaaaca tttcayacca cagaacgtga aatattcctt 60
taccatgtgc a 71
<210> 196
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 196
agctttggca gtttgctggt atatttttct gtatgrttat tgtagttaat ggtgacgtaa 60
ttttaccaac t 71
<210> 197
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 197
ctgacacttc acaaatgagg acagtacaga agttgktttt tttaacgatc attttaatct 60
atcattccta c 71
<210> 198
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 198
ttaggtgtaa tggtgtggtt aatgattggg ctgtgygtga aggctgtaaa atcccacata 60
aattacaaca t 71
<210> 199
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 199
ggccatgatc taaacccaag acgcacattt taaaakacaa aacttgtcca cagagaagtg 60
aggggaaaaa a 71
<210> 200
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 200
atgttgatgt ggagaaattc tgtggtcatg gcaacmgtcc tggttaaatc ctagaccaat 60
cacaaagcag a 71
<210> 201
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 201
gacacattgc tagtaattag attcttgact aaggtmtttt cagcaattct ttggttgtca 60
ctgtcaggtt g 71
<210> 202
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 202
gcttgctcaa aggcagaaaa gtggaggaaa aacaaktcaa ttatggtgcg cggcgtcgac 60
aaagacaggt g 71
<210> 203
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 203
atagttgcga tgttttaatt taaacctgtc atccaygagc agcagctaaa ctcattggca 60
tgtgtgtgtg a 71
<210> 204
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 204
atcttggtta aatatgagat cgtagaatta ccatamaacg tcactgtaca cctgttttga 60
tgcgctagaa g 71
<210> 205
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 205
atacttaggc ttattagtaa gaatcgtgtt attackcgaa tttaaccata ttcatttagc 60
acatcgcatt t 71
<210> 206
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 206
aggcagcgga tgtgaaatca tctgctctgt gtggaygtgc caattccaaa catttgagtc 60
tgtgcatatt t 71
<210> 207
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 207
taaaagaaat gctacagtat gtgactttta tgcacracat gaatttgtat tattgcactt 60
ctgggcatga g 71
<210> 208
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 208
gagagagaag atcaaagaca taaaagtaga tttcckatca gtccaaatga gtgacagagt 60
cttaagatac c 71
<210> 209
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 209
gtcatcgtct ctgatgcatt attcagactc actacmtcaa ccctagacag atcctatctc 60
aacagggaaa t 71
<210> 210
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 210
ctaaaagtaa atccaactcg acacagaggt tagtgrataa cgttttttct tgctgttgcg 60
atttccatat t 71
<210> 211
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 211
caaaacacag aaccgctgta agatgatagg agggtrttga cactgtatgc tgttgtgtca 60
cttacaaaca c 71
<210> 212
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 212
atcacagttt gcaaaataaa atgacatggc agatgktgga ctgtgtttaa actaaaacag 60
caccgtgggg a 71
<210> 213
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 213
taataattac tcctactgaa gaagtattca ttttgyatgt aatgcagatt tttcagctct 60
aattacagta a 71
<210> 214
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 214
cctaagcact ggccatgaag ggctgtgacc agtcaragcg tgttgtaatg ttgttgacgt 60
aatgtcactg t 71
<210> 215
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 215
gggagtctgt ccacttacaa ctcacctact gcaatkttta agaactagag tttttggatt 60
ggacagaggg c 71
<210> 216
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 216
agaaaaaagc ttagccgtgc gtgcagctct gaaacytagg tgccatcttc ctcgctcctc 60
actcttcctc t 71
<210> 217
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 217
ggtggagctc actgatgaac gctcagctct tcaacrggaa ctgacttctg tccaggaaac 60
ggtggctcgg g 71
<210> 218
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 218
gcagcagata atggccttgg caggtgttgc cagggraatg cccccagtgg tgtgagcttt 60
taatgaccac t 71
<210> 219
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 219
tctttgggaa actctacacg aagttgcacg ttcacrcggc ggaaaaaagt gaaacgaatc 60
cttgcgcttg t 71
<210> 220
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 220
ccctgaccgg tctaaacatc atactgagga agacgragac gaagactgac aatcaaaacg 60
tcaaaagaca a 71
<210> 221
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 221
atgattgctt gctgtgtaaa agcgcctgat gacgarcaca caatgcaacg atgatctgtt 60
attcatggca t 71
<210> 222
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 222
ttaatctcac ccattctttc tccatctccg ttcttmcttt ttctttttct atttgcacct 60
gcacgactcg t 71
<210> 225
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 225
atgtcgctgt ccaactcagc cacgccccca ggaagmtcga tgaccaccgt ggactgaaag 60
aatcaagtca a 71
<210> 224
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 224
gaagttcacc atatggacgc acatgactcc gttgcrtgcc gtgttttcat agttctaaat 60
ctagtcacaa g 71
<210> 225
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 225
agcagacaca ggaagtgggg cagaaacacc aaacakgggg ggcagggggg ggctcacctg 60
tcacagtgag g 71
<210> 226
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 226
aaactgcaga cacacacaag tctgctactc accgtraact cccgcagatt ttcacatttg 60
tgtggctcct c 71
<210> 227
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 227
tgcaaacaca catttagctg tgacactaaa atctargggc acaaaccgtc catcactgct 60
gcatagaaga a 71
<210> 228
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 228
atgacgatcg gagtcaaatg tgtgatgaaa agctgragca gggaggcaga aatgaaagga 60
ctgggtgaaa t 71
<210> 229
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 229
tcagactgag ctgaggagag tgtgtaagga gtatgrggtg tgtttccaag cctactcgtc 60
tttagggaga g 71
<210> 230
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 230
ttgttttgag caaattaagt aagaaatggg gcacgrcaac tgtttatagc accttgtata 60
ggacattgac a 71
<210> 231
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 231
agaaagtaag agttcagttt ggggctcaaa tcaagractt ttctatccat aaagtaatag 60
tgtatgattt c 71
<210> 232
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 232
cgtgaggaca gaggtttgaa agaaatgttc tgtggyccct gggttcgctg agagacatat 60
tgatcaatag a 71
<210> 233
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 233
agtcctggac agacgagaac atgctgtttt ggctcygtgt ttggctactt tttaatctga 60
tgtaatctgc t 71
<210> 234
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 234
agcgcttcag tgatatgatt ctgtgtgtca ataacyatta tggaaggatc gcgacttcaa 60
gccattgatg t 71
<210> 235
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 235
tatcctgaga ggcaatccct tcaatatttg tcaagraatt tccctaaaaa ttgtcatccc 60
gccaccggct g 71
<210> 236
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 236
ttccttttct atgtctctca ctctagtcct tgtttyaaga gctgcttcac ttacaatcat 60
catagactga t 71
<210> 237
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 237
tctttcctct gcagttctgg atgaaagagg gttgcrggct cttgttgcca ttaccaaata 60
tggaagaaat t 71
<210> 238
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 238
ccttaaaata aggtaacgac tcattaaaac caaackttca tgccattatt ttgacgttgt 60
tccacacaca c 71
<210> 239
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 239
ctcctccaac cttatccaga caagtgtgta catccygctg ctgtggttaa tgtccgtgtg 60
tgtgaatgtt g 71
<210> 240
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 240
ttccctttcc atctacagtg atatgagaaa caccaygact taacggtgca atactccctc 60
tgctggatgt t 71
<210> 241
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 241
acagttatga attacactga ctgaggggga gcaggrgctt cagaatcaat tcatcatcac 60
acaaatggaa a 71
<210> 242
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 242
tgtcaaatca ggtgacgaag aacgccaacg tcatcmaatc aaccaatcag atgcgtttaa 60
gggcggtatt t 71
<210> 243
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 243
gagcatcttg aatggagaga tagattcaac cttccraacg gcataaagag aaataagtat 60
caaaacatca c 71
<210> 244
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 244
taaaacatct ctttggtctg tcacaaacgc aaacaraaaa cgctgcactg attttgacaa 60
acgttacttc a 71
<210> 245
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 245
aggatcaagg tcttcttcca ttgaaggctg aagctragaa ctacgggaaa agttgtttcc 60
ttttctataa a 71
<210> 246
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 246
cgtgtctttt tctgccattt cacttttggg agagamatca tcataacaca caacaaggta 60
atggcacaaa c 71
<210> 247
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 247
acagcaggtc tggttgtctg ctccctcaat gcttcyttcc tcttttacac agcagtgaca 60
gcataagtag c 71
<210> 248
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 248
attttgtttt tgtgctttta ttttttagac tctctmattt ttgtttatga catttatcct 60
actgatggac a 71
<210> 249
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 249
gtgttatcta tctatctatt atcatcatca tcattrttgt tagtatttga ttttgtgttt 60
tctctttttg g 71
<210> 250
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 250
tataaaaatc acaaaattaa catttcaatg aaccartgat aagcaattaa aatacttttt 60
aactgaaaac a 71
<210> 253
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 253
ggtgatgagg caccggcaga cctccagtcg acaagragac catttacagc cattttacat 60
ctttaaccag t 71
<210> 252
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 252
gacagcagtg gtttggagag cagtcaaatt gatggrcagt ccagtccagg agcgtgtgtt 60
gaatcctctc c 71
<210> 253
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 253
gggctgccga ctcccgggct gcgttcatgg acacgkgttt tgatttgtgt ggttcaggac 60
atttacataa t 71
<210> 254
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 254
tgtcatgtca tctgtaggca acaaaccttc tccagyccca tgttatccag atacagctcc 60
ttcagtgacc t 71
<210> 255
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 255
agagattata tttcttatta aagaaagaag cagcgygttg ttatttagca tattcacagc 60
tgtgacaggt c 71
<210> 256
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 256
ccataactcg attagcacag tggaatgtat gacgtmcaac cagagaatgt aaacaaagcc 60
atcttctact t 71
<210> 257
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 257
tgttgactta gaatccagat gaggcagcat cacacrgacc agagtgttgg ctaaacccaa 60
agaccaggag a 71
<210> 258
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 258
tgttaacatg actgtttctt tctgttctcc cacagkgtta ccaatcatcc aggatgtctt 60
caaagcctca c 71
<210> 259
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 259
cagaagcgca gtgctgtgtt ggcagattta caagaytcca actggagaag acgtttagct 60
ctgtagttgg a 71
<210> 260
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 260
tcaacatgta atagcagcag gttgtctcag gttcartgtt aactaaggag tttatagatg 60
ctgtgaataa t 71
<210> 261
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 261
tatataggta actacagttt acactactaa cctacraatt cacttggctt cagtcacttc 60
tgttgaggtg t 71
<210> 262
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 262
cgcattcaca tccagtcgtg aaaaccctcc gacacrcgca tacatgcaaa gggcagcatc 60
ccagtaccac c 71
<210> 263
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 263
tgccgtggga aaagagtaca gtaaacctga cagacraggg ataaaacaca aaaattaaac 60
aacgtgaacg a 71
<210> 264
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 264
atatatgaga gcaaatcatt atcattgcca tcatcracaa aaacagcaaa ttatttaata 60
ttatggactt a 71
<210> 265
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 265
tttagaattt caaaagaaac ctctgaataa aacacmggtc tcagtctctc tcggtcgatt 60
cccctctctc t 71
<210> 266
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 266
ctgtaattgt acttatgagg ccaccaacgc tccagraagt gtagtccaca tgctttgaaa 60
acaaatcttt c 71
<210> 267
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 267
tgtgcagcgc tgctgttttt caataattca taaacratgt ggcttgaagc actttggtga 60
ggctgcaggg g 71
<210> 268
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 268
cctctgctgg ttcggtactc gtgaccacac tgtggygacg gtttcagatt ctggtctatt 60
ctaaacagag c 71
<210> 269
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 269
gatctgattg gtaggaatcg gttaggagac gctggyggtc ggcctgagag gttggaaaaa 60
gacaacagac a 71
<210> 270
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 270
atcagcaagt agagagcatc gtggttggag acatcrtcgc tgctccgtac agagaccacg 60
gaacctggaa c 71
<210> 271
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 271
aggtctgttt catcattccc aggggtgcct ccatgygctc acgatgctct gcttccttca 60
tacctagttc t 71
<210> 272
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 272
ataaaggact attgtatttg gtcataattt gtctcmagct ccttttgttg gtgataaagt 60
tgtattgtga a 71
<210> 273
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 273
aggtcatttt ccacagtgga gatcacaaac aaaagyttca gcatctaata atgagcatca 60
tcttattggc t 71
<210> 274
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 274
actggtatga ggggaaaagt ggtgaaggag gcagartgtt gaggaagata tttcggtagc 60
ctctcgcagt c 71
<210> 275
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 275
ggtctgagtg aaactgagaa acgcagagca aaagcktcag cgtgaaatct ttccaagaac 60
gatcgcagtc t 71
<210> 276
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 276
tggtaatgtt caatcatcac tgatatgcat ggatcygtaa tcaccagccg tcagtgttcc 60
tcaggaagtg c 71
<210> 277
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 277
gccctacatc cccctatcgt ctggcctcac tgtttytcat ctgcttgtgt caactatctt 60
tcccatattc t 71
<210> 278
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 278
ggttgctgtg ctcctgtata tcagatgtta atgccrggct ggaacacact gcactacatc 60
aagttacgtc a 71
<210> 279
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 279
atgattcaca ccatcaaata tttagtgcag agaaamgtcc aagggcatat gctgaaagct 60
aattccagct g 71
<210> 280
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 280
ttaataaacg cgttaactct tgttttcagt gtaggycttg acgtgacaaa actttcctat 60
actaccagcc t 71
<210> 281
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 281
ttctctagct ttaaagtttc ggccacaaac tacagmctat aattgacatc ctgaattcaa 60
aatccaaagt g 71
<210> 282
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 282
tagttcccaa ccaggtttga ctggcgatga cagacmtgga tggcagtcga tatgtcaagt 60
cggtcacttt t 71
<210> 283
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 283
tcttttttct tcttccactt cttttctgtt gcttamcgca ctgtgttcaa ctatctcaaa 60
cacatgcata c 71
<210> 284
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 284
caatggtgtc gctgtcatcc attttgctgt gtgctyggtg ccactcctca gacacccagg 60
caggttatgt c 71
<210> 285
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 285
tggaacttta atgctgtgac ttatctttcc tactgkggaa gtcaaaatgt ttgctgaaag 60
gatccgagaa t 71
<210> 286
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 286
agcctagcct ggtggtgtgg tgggatgtcg gttggrcgac ttgtgttttt atcaggctgt 60
caaaatgaat g 71
<210> 287
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 287
atcatgcggt gcaggtctgg tctctttagt acattmgcag agtcactttc tatggcaacc 60
cagtctcgca g 71
<210> 288
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 288
gaggattggt gatcagacag agggcttatt gtcacrcttg ttcgtagaca tggattaacg 60
ataccaggcc c 71
<210> 289
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 289
tctctgaaat aaaccttctc tatccttgaa cctctrttct tactgctatt accaccatca 60
ccactactgc t 71
<210> 290
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 290
tctaaacata aagtgacgct tcaagtggac tacacrcaaa cacagaaaca gaaagaatta 60
gggagagagg t 71
<210> 291
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 291
caaggtcaca cactctagag caaatacatg cagggyatat gaggttggct tgtacagtca 60
tatttgtaca c 71
<210> 292
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 292
aatgctgcac agttacaaca agtatgtgaa gaattrggaa gccggggcca ggactgtaag 60
aagaagtgaa g 71
<210> 293
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 293
gtttgtgact cggtccctat agcctctcat gatgtygtct aatatttcca cctgaaattc 60
tgagagataa g 71
<210> 294
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 294
accaaattag cttgattgca atacgttagg ctatgrtgct gttttttaat ggtaatggtc 60
ataaacagtg a 71
<210> 295
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 295
gttgtgtccg ggacaggacg ccggcgtcag ctccakgtct ggcggtgatc cctgcatttt 60
taaacagctg c 71
<210> 296
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 296
ttaggcctgg atcagtacca gaaccgtttc agaacrtttg gcacataaac aaacaacacg 60
acacattttc g 71
<210> 297
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 297
tcttagtaga cgttcccagg tgtgagtgag tgcaarcgca cagtcttatg catacacaat 60
cacacaaaca g 71
<210> 298
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 298
ttcctgtatt tttgtcatat cacctgaaat tactgmtatt cacaggctct acatccgtcc 60
tttactctgc t 71
<210> 299
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 299
caacacgaca aaggctcatt gactgttaat aggacrcaat ttcaatctgt aaaatctgta 60
tttatagcca c 71
<210> 300
<211> 71
<212> DNA
<213> Cynoglossus semilaevis (Cynoglossus semilaevis)
<400> 300
gtaactccag aaacatacca aaaccagaca gcaacycttc agaagtttca gaatttgagt 60
tcagacagga t 71
Claims (7)
1. The SNP locus related to disease resistance of cynoglossus semilaevis is characterized in that the SNP locus is a nucleotide sequence shown in SEQ ID NO: 1 to SEQ ID NO.300 at position 36.
2. The application of the SNP locus of claim 1 in preparing a detection product for breeding of a disease-resistant improved cynoglossus semilaevis variety.
3. The use of claim 2, wherein the assay article is a gene chip.
4. A gene chip, which is used for detecting the SNP locus related to the disease resistance of cynoglossus semilaevis as claimed in claim 1.
5. The application of the SNP locus of claim 1 in screening of cynoglossus semilaevis disease-resistant individuals.
6. A method for screening disease-resistant individuals of cynoglossus semilaevis is characterized in that the method is to use the gene chip of claim 4 to perform genotyping, calculate the breeding value GEBV of an individual genome and screen out individuals with disease-resistant potential according to the size of the GEBV.
7. The method of claim 6, wherein the method comprises the steps of:
1) establishing a reference population for disease-resistant breeding of cynoglossus semilaevis, evaluating the accuracy of the optimal linear unbiased prediction GBLUP estimated breeding value of the genome by using the reference population, and comparing the accuracy with the prediction accuracy of the optimal linear unbiased prediction ABLUP based on a pedigree;
2) extracting the genome DNA of the candidate parent fish of cynoglossus semilaevis and carrying out genotyping by using the gene chip of claim 4;
3) estimating a genome estimated breeding value GEBV of the candidate parent fish by using the constructed cynoglossus semilaevis reference group and the genotyping information of the candidate parent fish;
4) and selecting individuals with disease-resistant potential according to the GEBV of the candidate individuals to cultivate the disease-resistant strains.
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| WO2023103303A1 (en) * | 2021-12-06 | 2023-06-15 | 中国水产科学研究院黄海水产研究所 | Genomic selection method for plectropomus leopardus disease-resistant improved variety breeding |
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| CN108084261A (en) * | 2017-12-29 | 2018-05-29 | 中国水产科学研究院黄海水产研究所 | A kind of Cynoglossus semilaevis anti pathologic immunity related gene and its application |
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| CN114990243A (en) * | 2022-06-21 | 2022-09-02 | 中国水产科学研究院黄海水产研究所 | Method for screening marker combination of anti-vibriosis harveyi cynoglossus semilaevis and screening disease-resistant individual of cynoglossus semilaevis |
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