CN105603065A - SNP locus related to litter size on sow chromosome 3 and primers thereof - Google Patents
SNP locus related to litter size on sow chromosome 3 and primers thereof Download PDFInfo
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- 210000000349 chromosome Anatomy 0.000 title claims abstract description 6
- 108700028369 Alleles Proteins 0.000 claims description 5
- 239000003147 molecular marker Substances 0.000 claims description 3
- 101100240528 Caenorhabditis elegans nhr-23 gene Proteins 0.000 abstract description 8
- 108090000623 proteins and genes Proteins 0.000 abstract description 8
- 230000009286 beneficial effect Effects 0.000 abstract description 2
- 238000012214 genetic breeding Methods 0.000 abstract description 2
- 230000002068 genetic effect Effects 0.000 abstract description 2
- 239000003550 marker Substances 0.000 abstract description 2
- 101150111062 C gene Proteins 0.000 abstract 2
- 101150050863 T gene Proteins 0.000 abstract 1
- 238000012163 sequencing technique Methods 0.000 abstract 1
- 241000282898 Sus scrofa Species 0.000 description 40
- 238000006243 chemical reaction Methods 0.000 description 7
- 239000003153 chemical reaction reagent Substances 0.000 description 7
- 238000009396 hybridization Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 241000282887 Suidae Species 0.000 description 4
- 230000001850 reproductive effect Effects 0.000 description 3
- ZHNUHDYFZUAESO-UHFFFAOYSA-N Formamide Chemical compound NC=O ZHNUHDYFZUAESO-UHFFFAOYSA-N 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- 241001237732 Microtia Species 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 238000009395 breeding Methods 0.000 description 2
- 230000001488 breeding effect Effects 0.000 description 2
- 230000004087 circulation Effects 0.000 description 2
- 238000010219 correlation analysis Methods 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000003205 genotyping method Methods 0.000 description 2
- PMHURSZHKKJGBM-UHFFFAOYSA-N isoxaben Chemical compound O1N=C(C(C)(CC)CC)C=C1NC(=O)C1=C(OC)C=CC=C1OC PMHURSZHKKJGBM-UHFFFAOYSA-N 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 238000012935 Averaging Methods 0.000 description 1
- 101100433727 Caenorhabditis elegans got-1.2 gene Proteins 0.000 description 1
- 108010007577 Exodeoxyribonuclease I Proteins 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000009027 insemination Effects 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 230000013011 mating Effects 0.000 description 1
- 238000000034 method Methods 0.000 description 1
- 230000000869 mutational effect Effects 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000012264 purified product Substances 0.000 description 1
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- 241000894007 species Species 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
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Abstract
The invention belongs to the technical field of biology, and relates to an SNP locus related to sow litter size characters, in particular to an SNP locus related to the litter size on the sow chromosome 3 and primers thereof. The SNP locus is Chr3:59537592 of a genome version NCBI Build Sscrofa 10.2. The allelic genes of the locus are T and C, and the locus has three gene types of C/C, T/C and T/T. The primer for amplifying the locus and the extending primer are Chr3-1-PCRU: 5'TTG GTC CTC AGT CAT GTG G-3'; Chr3-1-PCRL: 5'AAC CTC TTC CCT TGA CCT GA-3'; Chr3-1-SNPU: 5'-TGA CTA GTA GGC AGG CAA GAA CAT TCA GAG-3'. The sow litter size of the T/C gene type is larger than the sow litter size of the C/C gene type, wherein T is the beneficial allelic gene of the sow litter size, and the T/C hybrid litter size is the largest. The SNP locus has the advantages that the whole genome re-sequencing method can rapidly screen out the SNP locus or genes related to the characters in batches, the SNP locus which is the Chr3:59537592 of a genome version NCBI Build Sscrofa 10.2 can be used as a genetic marker and used for improving the sow litter size for sow genetic breeding.
Description
Technical field:
The present invention relates to the SNP site relevant to pig number born character, be specifically related on No. 3 chromosomes of pig oneIndividual SNP site and the primer thereof relevant to nest litter size, belong to biological technical field.
Background technology:
The characters of number born of pig is the chief component of pig reproductive trait, improves litter size of pig to improving pig industryOverall economic benefit significant. The Taihu Lake pig of China is well-known with high yield, nest litter size than great majority otherChinese native pig breed and abroad many 4-5 of nest litter size head of pig kind (" Chinese pig breeds will " Zhang Zhongge chief editor,Science and Technology of Shanghai publishing house, 1986). The polymorphic site relevant to high litter size in the pig of Taihu Lake, can serve as heredityMark is for improving the litter size of sow.
The heavy sequence measurement of full genome has been widely used in the correlative study of each species, and it can be fast all sidedlyFilter out and proterties, gene or mutational site that function is relevant, made up QTL location and candidate gene sideMethod wastes time and energy, the deficiency such as single.
There is not yet the open source literature report that in the present invention, SNP site is relevant to Litter Size in Pigs at home and abroad.
Summary of the invention:
The object of this invention is to provide on No. 3 chromosomes of pig a SNP site relevant to nest litter size andPrimer.
The present invention has selected Erhualian (one of Taihu Lake pig), hides pig, the fragrant pig of bar horse, Laiwu Pigs, and YunnanSouth 5 Chinese native pig breeds of microtia pig and wild boar have carried out the full genome order of resurveying. The order of resurveying of 6 pig kindsNumber of individuals is respectively 11, and 11,9,10,10 and 10. The order sequenced data of resurveying is followed with reference to baseCompare because of group (NCBIBuildSscrofa10.2), utilize SAMTools software to carry out SNPs siteExtraction. Inventor has compared Erhualian and other 5 pig kinds, finds a SNP on No. 3 chromosomesThe litter size phase of site (Chr3:59537592 of genome version NCBIBuildSscrofa10.2) and pigClose. The allele in this site is T and C, has tri-kinds of genotype of C/C, T/C and T/T.
Screen behind this site, inventor is at white Du Luoke × Erhualian hybridization resource population F2Godmother pigIn body, it is verified and correlation analysis, found that this site and litter size of pig have significant correlation.
Beneficial effect of the present invention is: the heavy sequence measurement of full genome can screen and proterties fast, in bulkRelevant SNP site or gene, the Chr3:59537592 of genome version NCBIBuildSscrofa10.2The litter size of genetic marker for the genetic breeding raising sow of pig can be served as in this SNP site.
Brief description of the drawings:
Fig. 1 is white Du Luoke × Erhualian F23 kinds, Chr3:59537592 site in godmother pig resource populationGenotypic litter size comparison (two kinds of genotype litter sizes relatively time in figure, * represent 0.01≤P≤0.05; * represents P < 0.01).
Specific embodiments:
(1) the population distribution feature in SNP site
The present invention selects Erhualian (one of Taihu Lake pig), hides pig, the fragrant pig of bar horse, Laiwu Pigs, and the southern regions of the Yunnan Province5 Chinese native pig breeds of microtia pig and wild boar have carried out the full genome order of resurveying. The order of resurveying of 6 pig kindsBody number is respectively 11, and 11,9,10,10 and 10. The order sequenced data of resurveying is followed with reference to geneGroup (NCBIBuildSscrofa10.2) is compared, and utilizes SAMTools software to carry out SNPs siteExtraction. Inventor has compared Erhualian and other 5 pig kinds, finds a SNP on No. 3 chromosomesThe litter size of site (Chr3:59537592 of genome version NCBIBuildSscrofa10.2) and pigRelevant. From the analysis result of the order of resurveying, this site equipotential base in other 4 Chinese native pig breeds and wild boarBecause the frequency averaging of C is 84.2%, and the frequency of allele T is 100% in Erhualian.
(2) qualification in SNP site
At design of primers website design SNAPSHOT primer, comprise one couple of PCR amplimer (Chr3-1-PCRU:5’-TTGGTCCTCAGTCATGTGG-3’；Chr3-1-PCRL：5’-AACCTCTTCCCTTGACCTGA-3 ') and extension primer (Chr3-1-SNPU:5 '-TGACTAGTAGGCAGGCAAGAACATTCAGAG-3 '). Totally 24 (12 pairs) primers in this site and other 11 sites are respectively got 1 μ l and are mixed into newlyMultiple PCR primer pond. First carry out 12 heavy PCR reactions, reaction system agents useful for same is: multiplex PCRPrimer pond (6.25 μ Meach) 8 μ l, dNTP (10mMeach) 8 μ l, 10 × PCRBufferII116 μ l,MgCl2(25mMStock) (5U/ μ is 24 μ l and ddH l) for 234 μ l, FastStartTaq2O460 μ l, will(50ng/ μ l) is mixed into 10 μ l systems and carries out 12 heavy PCR for above reagent mixed liquor 8.5 μ l and 1.5 μ lDNAAmplified reaction. Amplification reaction condition is: 94 DEG C of 4min; 94 DEG C of 30s, 55 DEG C of 30s, 72 DEG C of 1min, altogether40 circulations. Next step carries out PCR reaction purification, and required reagent is: ExoI (10U/ μ l) 60 μ l,(1U/ μ is 130 μ l, 10 × SAPBuffer35 μ l and ddH l) for SAP2O125 μ l, at the PCR of each sampleAfter adding the above-mentioned purifying mixed liquor of 3.5 μ l centrifugal in product, react by following program: 37 DEG C of 40min, 96DEG C 10min. Then carry out single base extension, required reagent is: SNAPSHOTMultiplexreadyReactionMix100 μ l, extension primer pond (6.25 μ Meach, 12 are extended primers and respectively get 1 μ l mixing)8 μ l, 5 × SequencingBuffer200 μ l and ddH2O300 μ l, by above reagent mixed liquor 6 μ lBe mixed into 10 μ l systems with the purified product of 4 μ l and carry out extension. Extension condition is: 96 DEG C of 10s,50 DEG C of 5s, 60 DEG C of 30s, totally 25 circulations. Then the purifying that carries out extension product, required reagent is:(1U/ μ is 120 μ l, 10 × SAPBuffer70 μ l and ddH l) for SAP2O140 μ l, in the extension of each sampleAfter adding above-mentioned 3.3 μ l purifying mixed liquors centrifugal in product, react by following program: 37 DEG C of 40min,75 DEG C of 20min, 4 DEG C save backup. The SNAPSHOT reaction that each sample is got after the above-mentioned purifying of 5-4 μ l is producedHigh-purity formamide (Hi-DiFormamide) of thing and 5-6 μ l is mixed into 10 μ l systems and is splined on ABI after centrifugal3730XL sequenator carries out Genotyping, uses GeneMarker software analysis somatotype result.
(note: the reaction reagent mixed liquor consumption of above 4 steps is 100 required amount of reagent of sample, from single alkaliBase extension starts, and needs lucifuge to carry out. )
(3) white Du Luoke × Erhualian reproductive performance hybridization resource population builds
Follow-up confirmatory experiment resource population individual sample used and proterties data thereof are given birth to by Agricultural University Of Jiangxi animalThing technology National Key Laboratory cultivation base provides.
In December, 1998, purchase in Erhualian conservation field, 3, Jiang Cong Jiangsu Province, Agricultural University Of Jiangxi's scientific research pig farmThe Erhualian sow of buying carries out the intersection breeding of blood relationship between field. January calendar year 2001 is according to parent's Farrowing Traits, from 3In the pure breeding offspring of boar and 11 sows, select female for generations as resource colony of 17 Erhualian sowsThis. 2 high-quality white Duroc boars that SygenPIC company is so kind as to give are as the male parent for generations of resource family.By these 2 white Du Luoke and 17 F that Erhualian hybridization produces1Breed for individuality, avoid complete sameBorn of the same parents' mating, and every boar conventionally with same insemination of sows, to obtain the F of large sample2Family half sibs.In January, 2003, March and June, project team is respectively by the 1st crowd and the 2nd crowd of F2It is suitable that sow is sent to Jiangxi ProvinceSpring city's herding seed multiplication farm (59), state-run Red Star kind pig farm, Jiangxi Province (52) and Jiangxi Shangyou County quaternionic breedingPig farm (118) is for the mensuration of sow reproductive trait.
(4) SNP site is at hybridization resource population F2The Genotyping of godmother swinery body
According to method above-mentioned steps (2) Suo Shu, with 192 white Du Luoke × Erhualian hybridization resource populationF2The individual DNA of godmother pig is the genotype in this SNP site of template detection. 192 F2For having in resource population individuality176 successful somatotypes of individuality, 3 kinds of genotype C/C, the number of individuals of T/C and T/T is respectively 60,84 Hes 32。
(5) regulating effect of SNP site to characters of number born
At F2In resource population individuality, be 10.5507 with the average litter size of C/C genotype individuality, standardDifference is 0.7672; Be 11.595 with the average litter size of T/C genotype individuality, standard deviation is 0.7568;Be 10.9155 with the average litter size of T/T genotype individuality, standard deviation is 0.8394.
The present invention adopts general linear model (GLM) to analyze SNP site to the total litter size of pig, product young number aliveImpact. All statistical analysis adopts SAS software to carry out. Model is as follows:
Yijklm＝u+Ai+Gj+Bk+Pl+eijklm
Wherein YijklmFor the total litter size of pig or product young number alive, u is colony's average, AiBe i individual additivityEffect, GjBe the fixed effect of j SNP loci gene type, BkFor a batch effect (k=1,2,3,4), PlFor stochastic effects (l=1,2,3).
Analysis result finds that there is significant correlation in this SNP site of Chr3:59537592 with litter size. With T/CGenotypic F2The litter size of godmother pig is than with average many 1.04 (P=0.0181) of the genotypic sow of C/C,As can be seen from Figure 1, minimum with the genotypic number born of sow of C/C. T is the useful equipotential that improves litter size of pigGene and T/C heterozygosis litter size of pig are the highest.
Find out from SNP site and litter size correlation analysis, Chr3:59537592 can do in this SNP siteFor being applied to molecular marker assisted selection (MAS), molecular labeling improves the litter size of sow.
Claims (2)
1. on No. 3 chromosomes of pig, a SNP site relevant to litter size is assorted at white Du Luoke × ErhualianHand over resource population F2Application in godmother's litter size of pig molecular marker assisted selection, is characterized in that: described SNPThe allele in site is T and C, has tri-kinds of genotype of C/C, T/C and T/T, genotypic with C/CNumber born of sow is minimum, and T is that useful allele and the T/C heterozygosis litter size of pig of raising litter size of pig is the highest.
2. a test right requires the primer in SNP site described in 1 assorted at white Du Luoke × ErhualianHand over resource population F2Application in godmother's litter size of pig molecular marker assisted selection, is characterized in that: described primerNucleotide sequence as follows:
Chr3-1-PCRU：5’-TTGGTCCTCAGTCATGTGG-3’；
Chr3-1-PCRL：5’-AACCTCTTCCCTTGACCTGA-3’；
Chr3-1-SNPU:5 '-TGACTAGTAGGCAGGCAAGAACATTCAGAG-3 ', described inSNP site is minimum with the genotypic number born of sow of C/C, and T is the useful allele that improves litter size of pigAnd T/C heterozygosis litter size of pig is the highest.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106399524A (en) * | 2016-10-11 | 2017-02-15 | 贵州大学 | PAV3 molecular marker related to litter size of Xiang pig and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2007133075A2 (en) * | 2006-05-12 | 2007-11-22 | Institute For Pig Genetics B.V. | Igf2 marker assisted selection for porcine reproductive traits |
| CN103757003A (en) * | 2013-12-26 | 2014-04-30 | 中国科学院昆明动物研究所 | Litter size related SNP (Single Nucleotide Polymorphism) locus of swine 13# chromosome and primers thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007133075A2 (en) * | 2006-05-12 | 2007-11-22 | Institute For Pig Genetics B.V. | Igf2 marker assisted selection for porcine reproductive traits |
| CN103757003A (en) * | 2013-12-26 | 2014-04-30 | 中国科学院昆明动物研究所 | Litter size related SNP (Single Nucleotide Polymorphism) locus of swine 13# chromosome and primers thereof |
Non-Patent Citations (3)
| Title |
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| PEKKA UIMARI ET AL.: "Whole-genome SNP association analysis of reproduction traits in the Finnish Landrace pig breed", 《GENETICS SELECTION EVOLUTION》 * |
| 杨怀谷等: "利用选择性清除方法鉴别影响太湖猪（二花脸、梅山猪）及野猪产仔数差异的SNP位点", 《广东农业科学》 * |
| 林国珊: "二花脸猪特异选择位点与猪产仔数的关联性分析", 《中国优秀硕士学位论文全文数据库（电子期刊）》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106399524A (en) * | 2016-10-11 | 2017-02-15 | 贵州大学 | PAV3 molecular marker related to litter size of Xiang pig and application thereof |
| CN106399524B (en) * | 2016-10-11 | 2019-11-19 | 贵州大学 | One kind PAV3 molecular labeling relevant to fragrant litter size of pig and its application |
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