CN107937557A - One and the relevant SNP site of pig economic characters and its application - Google Patents
One and the relevant SNP site of pig economic characters and its application Download PDFInfo
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Abstract
The present invention relates to SNP site, specifically discloses one and the relevant SNP site (Chr7 of the multiple important economical traits of pig：34913640) and its apply.Multiple important economical traits of the invention by measuring and recording duroc, GWAS researchs are carried out to 3702 duroc bodies using the GBS technologies of optimization, have obtained a SNP (Chr7 significantly correlated with the multiple important economical traits of pig (thickness of backfat, eye muscle area and lean meat percentage)：34913640) site.Count the SNP site and resurvey the local pig breed (Jinhua Pigs, Wuzhi Mountain pig, Luchuan pigs, plum mountain pig, Rongchang Pig, Laiwu Pigs, Erhualian, river bend pig and people pig) of sequence and the SNP frequencies in market pig kind (duroc, Landrace, Yorkshire and Ternary Pig pig), it was found that its SNP frequency distribution is in local pig breed and market pig kind, there are significant difference, G is advantage allele in market pig, T is advantage allele in Native Pig, and scientific basis is provided for the marker assisted selection of pig.
Description
Technical field
The present invention relates to SNP site, specifically, is related to one and the relevant SNP site of the multiple important economical traits of pig
And its application.
Background technology
China is a pig raising big country, and the market demand of pork yield and quality increasingly increases, and improves pork yield, improves
Pork carcase quality, becomes the work that breeding scientist constantly explores for a long time.The breeding work of early stage focuses primarily upon pig
Phenotypic Selection, continuous with genome work promote and the extensive exploitation of genetic marker, and molecule selection is increasingly becoming can
Lean on and effective system of selection.
Single nucleotide polymorphism (single nucleotide polymorphism, SNP) mark is third generation SNP
Point, refers to a kind of polymorphism for having the mutation of single base on genomic dna sequence and producing, and this mutation includes single base
Transversion, conversion, insertion and missing.The SNP amounts of having the advantages that are big, high-frequency, low mutation rate, are widely used in gene component
Analysis, biological information automatic detection, simple and complex disease genetic research, herding breeding mark and global racial inheritance etc.
Research.SNP site assisted selection, is that objective trait is made choice on a molecular scale, can not be affected by environment, is led to
Genetic background selection is crossed, Linkage drag is reduced, so as to accelerate breeding process and precision.
Whole-genome association (Genome-wide association studies, GWAS) is livestock and poultry economic characters
Genetic improvement and the important method of mechanism parsing.With the development of two generation sequencing technologies, full-length genome resurveys sequence and simplifies gene
Group sequencing technologies become the powerful of high throughput SNP partings, and GBS (Genotyping-by-sequencing) is simplified gene
The classical representative of group sequencing, is a kind of efficient full-length genome SNP classifying methods, Direct Identification SNP can go forward side by side from colony
Row parting, tens of thousands of SNP parting information not waited to hundreds of thousands can be obtained with relatively low cost, have been widely used in animals and plants
(De Donato in the researchs such as SNP site exploitation, population genetic analysis, whole-genome association and genome selection and use
Et al., 2013；Elshire et al., 2011；He et al., 2014).
How much the thickness of backfat of pig represents fat, and back-fat thickness is thicker, and lean meat percentage is lower, on the contrary, then lean meat percentage is high.Lean meat percentage
Refer to the cutability that pig contains, the ratio shared by lean meat, the lean meat percentage of breeding pig can be higher.Pig eye muscle area refers to pig eye muscle
Cross sectional area, the lean meat percentage of pig individual is higher, and eye muscle area is bigger.Three above character has strong phase with domestic pig meat production
Guan Xing, has important research significance in breeding.
If a kind of and above three economic characters significantly correlated SNP site or SNP marker can be screened, help
In providing favourable theoretical foundation for the marker assisted selection breeding of pig.
The content of the invention
In order to solve the problems in the existing technology, the object of the present invention is to provide one and the multiple Important Economics of pig
The relevant SNP site of shape and its application.
In order to achieve the object of the present invention, technical scheme is as follows：
On the one hand, the present invention provides a kind of and relevant SNP site of pig economic characters, which is genome version
The Chr7 of Ensembl Sscrofa 10.2：34913640, the allele in the site is G and T, there is tri- kinds of G/G, T/T and G/T
Genotype.
Multiple important economical traits of the invention by measuring and recording duroc, carry out 3757 Duroc boars
GBS is sequenced, and identifies the whole gene group that 102,254 SNP cover pig, and remaining 66,737 are used for 3702 after strict quality control
The GWAS researchs of the multiple important economical traits of the head purebred colony of duroc, have obtained one and the multiple important economical traits of pig
(thickness of backfat, eye muscle area and lean meat percentage) significantly correlated SNP (Chr7：34913640) site.The SNP site is counted to resurvey
Local pig breed (Jinhua Pigs, Wuzhi Mountain pig, Luchuan pigs, plum mountain pig, Rongchang Pig, Laiwu Pigs, Erhualian, river bend pig and the people of sequence
Pig) with the SNP frequencies in market pig kind (duroc, Landrace, Yorkshire and Ternary Pig pig), find its SNP frequency point
Cloth is there are significant difference in local pig breed and market pig kind, and G is advantage allele in market pig, and T is advantage in Native Pig
Allele.
Wherein, market pig shows as that faster than Native Pig weightening, lean meat percentage is high.
Further, the economic characters are the thickness of backfat, eye muscle area and/or lean meat percentage.The equipotential of i.e. described SNP site
Gene and above three economic characters are significantly correlated.
The SNP site is located at the 101st bit base in the nucleotide sequence shown in SEQ ID NO.1.
On the other hand, the application the present invention provides the SNP site in the marker assisted selection breeding of pig.
The application is embodied as a kind of method for carrying out assistant breeding to pig using the SNP site, it may include with
Lower step：
(1) genotype of the sample pig in the SNP site is detected；
(2) sample pig of the selection with advantage allele genotype carries out the selection and breeding of advantage strain.
Preferably, selection genotype carries out the selection and breeding of advantage strain for pig homozygous G/G.
Wherein, the advantage strain is mainly shown as lean meat percentage height.
Further, the step (1) can use direct Sequencing, or first expand the fragment containing the SNP site and detect again
Mode carry out, for example, design primer, amplify the piece containing the SNP site from the sequence shown in SEQ ID No.1
Section, then detect the allele on the site.
Present invention also offers application of the SNP site in market pig/Native Pig economic characters advantage strain is identified.
On the other hand, the protection of the present invention is fallen within for expanding the primer pair containing SNP site fragment of the present invention
Scope.The primer pair is act as：The fragment containing the SNP site is amplified from the sequence shown in SEQ ID No.1.
The beneficial effects of the present invention are：
The present invention is to pig Chr7：34913640 SNP site carries out Genotyping, and multiple heavy with pig to the SNP site
Economic characters (thickness of backfat, eye muscle area and lean meat percentage) are wanted to be associated analysis, analysis is found, SNP frequency distribution is in Native Pig
Kind and market pig kind in there are significant difference, G is advantage allele in market pig, and T is advantage allele in Native Pig, is
The marker assisted selection of pig provides scientific basis.
Brief description of the drawings
Fig. 1 pig 100kg weight thickness of backfats (A), 100kg weight eye muscle area (B) and 100kg weight lean meat percentage (C) GWAS
As a result Manhattan figure.
Embodiment
Following embodiments are used to illustrate the present invention, but are not limited to the scope of the present invention.
1 pig economic characters whole-genome association of embodiment
1st, test material
Using the purebred colony of duroc as research object, 33,960 to be born in August, 2007 in January, 2016 are collected
The production performance record of body (12,987 boars and 20,973 sows) is used for the present invention.Traditional performance is measured in strict accordance with pig
Field internal specification carries out, including the 100kg weight thickness of backfat (Back fat thickness at 100kg live weight,
BF), 100kg weight eye muscle area (Loin muscle area at 100kg live weight, LMA) and 100kg weight are thin
Meat rate (Lean meat percentage at 100kg live weight, LMP).
2nd, test method
2.1 conventional determining characters
The 100kg weight thickness of backfat, eye muscle area, lean meat percentage：When pig weight reaches 100 ± 5kg, end is proceeded by
Survey；Using the thickness of backfat and eye muscle area at Aloka SSD-500 type B ultrasound instrument measure inverse 3-4 intercostals, and estimate lean meat percentage.
Probe, probe mould and tested position should be close during measurement, but should not weight；Probe straight line plane is hung down with pig back of the body median line longitudinal axis face
Directly, can not chamfer；When identifying ultrasonograph, it is first determined connective tissue and longissimus dorsi muscle sarolemma institute between skin interface, fat
The strong echo shadow bands produced, it is then determined that the strong echo image of eye muscle surrounding sarolemma, to determine eye muscle area shadow bands.
The 2.2 pig full-length genome SNP classifying methods based on GBS technologies
By simulating digestion pig genome, 36 kinds of common II type restriction enzymes and 24 kinds of double digestion groups are predicted
Close the digestion effect in pig genome；According to research purpose and colony's feature, I-Msp of EcoR, I double digestion combinations are selected to be used for
The GBS of pig builds storehouse, and GBS experiments and analysis process are optimized, and establishes the pig full-length genome SNP based on GBS technologies
Classifying method.By carrying out GBS sequencings, 102 identified to 3757 durocs of Wen Shi Durocs, 254 SNP are covered
The whole gene group of pig.
2.3 whole-genome association
To the 100kg weight thickness of backfat (BF), 100kg weight eye muscle area (LMA), 100kg weight of duroc colony
Totally 3 characters have carried out whole-genome association to lean meat percentage (LMP).
2.4SNP Quality Control
In order to obtain reliable GWAS as a result, the present invention carries out Quality Control using the following conditions：(1)MAF≥0.05；(2)HWE
≥10E-6；(3) two kinds of homozygotic individual numbers of each SNP are >=30.
2.5 with the significantly correlated SNP site of economic characters
The detection in the notable site of genomic level, Bonferroni corrections, independent marking number are carried out using independent marking number
Calculating using PLINK indep-pairwise orders obtain, obtain the p of 5% level of signifiance of Bonferroni genomic levels
It is worth for 0.05/14,084=3.55 × 10-6, and potential associated p value threshold value is 1.0/14,084=7.10 × 105。
According to the standard, 5% genomic level for reaching Bonferroni corrections with pig diversified economy character is obtained
Significant SNP site (P<10-5.45)。
3rd, result and analysis
The present invention utilizes 102,254 SNP pairs of the GBS sequencing acquisitions of optimization using 3702 Duroc colonies as object
3 important characters of pig have carried out GWAS analyses, it is determined that one and the pig 100kg weight thickness of backfat, 100kg weight eye muscle areas
Significantly correlated SNP (the Chr7 with three production traits of 100kg weight lean meat percentage：34913640), as shown in Figure 1.
Embodiment 2SNP (Chr7：34913640) frequency distribution in different cultivars pig
1st, test material
Local pig breed：6 Laiwu Pigs, 5 Erhualians, 6 river bend pigs, 6 people pigs, 5 Jinhua Pigs, 6 Wuzhi Mountain
Pig, 6 Luchuan pigs, 14 plum mountain pigs and 9 Rongchang Pigs.Market pig kind includes：16 durocs, 12 Landraces, 8 about
Gram summer pig and 36 Ternary Pig pigs.
2nd, test method
The extraction of 2.1 genomic DNAs
Genomic DNA is extracted using the GIAamp DNA Mini kits of QIAGEN companies, concrete operation step is as follows：
(1) the ATL buffer solutions of 180 μ l are added into the centrifuge tube of 1.5ml, add 20 μ l Proteinase Ks, are mixed；
(2) the ear tissue sample of 20mg or so is taken to be put into above-mentioned solution, 55 DEG C of digestion 8h；
(3) the RNase A of 3 μ l are added into the tissue fluid digested, 30min, purpose are placed in 37 DEG C of thermostat water baths
It is the RNA in degraded tissue solution；
(4) 200 μ l AL solution are added into solution again, vortex vibration is put into 70 DEG C of water-baths after mixing to be digested
10min；Treat that centrifuge tube recovers to room temperature, add 200 μ l absolute ethyl alcohols, be vortexed again for concussion and mix；
(5) above-mentioned solution is all added in DNA adsorption columns, after room temperature places 2min, 13000rpm centrifugations 1min；
(6) after centrifuging, filter liquor is abandoned, and adsorption column is put into another new 2ml collecting pipe, adds 500 μ l
PW1 solution, 13000rpm centrifugations 1min；
(7) after centrifuging, then adsorption column put into another new 2ml collecting pipe, adds 500 μ l PW2 solution,
13000rpm centrifuges 3min；
(8) solution in collecting pipe is outwelled, the liquid of the collecting pipe mouth of pipe is cleaned with paper handkerchief, adsorption column is placed on collection again
Guan Zhong, 13000rpm centrifuge 2min；
(9) lid of DNA adsorption columns is opened, be put into the 1.5ml centrifuge tubes of the number of finishing, room temperature places 2min, makes
Remaining ethanol is evaporated completely in pipe；
(10) to adsorption column center add 120-150 μ l AE buffer solutions, room temperature place 2min after, 13000rpm from
Heart 1min, resulting solution are genomic DNA.Genomic DNA is used after the detection of 1% agarose gel electrophoresis is qualified
NanoDrop quantitative concentrations；Concentration is uniformly diluted to 50ng/ μ l again, for testing in next step.
2.2SNP(Chr7：34913640) frequency in each breeding pig full-length genome weight sequencing data
Send each breeding pig genomic DNA to sequencing, statistics SNP (Chr7：34913640) in above local pig breed and commodity
Frequency distribution in pig kind weight sequencing data.
3rd, result and analysis
SNP(Chr7：34913640) such as institute of table 1 of the SNP frequency distribution result in different places pig kind and market pig kind
Show, there are significant difference in local pig breed and market pig kind.G is advantage allele in market pig, and T is advantage in Native Pig
Allele.
Table 1SNP (Chr7：34913640) the SNP frequencies in different places pig kind and market pig kind
A kind of and relevant SNP marker of the multiple economic characters of pig has been obtained, in the low colony of lean meat percentage, has passed through selection etc.
The individual of position gene G carries out breeding, can improve the lean meat percentage of colony after breeding.
Although above the present invention is described in detail with a general description of the specific embodiments,
On the basis of the present invention, it can be made some modifications or improvements, this will be apparent to those skilled in the art.Cause
This, these modifications or improvements, belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Sequence table
<110>China Agricultural University
<120>One and the relevant SNP site of pig economic characters and its application
<130> KHP171115705.8
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 201
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 1
atcagaaacc aaacaaaagc taagtccctg gtgcctcagc tggttaagga tctggcattg 60
tcactgcagg ggcttgggtc actgctgcgg catgggtcac tctgaattct atggggagcc 120
acagaagaaa gacatagacc ctaactcagg atctctttct ctcctcctgt caatgggaga 180
cgagaggacg tgaaagcttg g 201
Claims (10)
1. a kind of and relevant SNP site of pig economic characters, it is characterised in that the site is genome version Ensembl
The Chr7 of Sscrofa 10.2：34913640, the allele in the site is G and T, and the economic characters are the thickness of backfat, eye muscle
Area and/or lean meat percentage.
2. SNP site according to claim 1, it is characterised in that G is advantage allele in market pig, T in Native Pig
For advantage allele.
3. SNP site according to claim 1 or 2, it is characterised in that the SNP site is located at shown in SEQ ID NO.1
Nucleotide sequence in the 101st bit base.
4. application of claims 1 to 3 any one of them SNP site in the marker assisted selection breeding of pig.
5. application according to claim 4, it is characterised in that comprise the following steps：
(1) genotype of the sample pig in the SNP site is detected；
(2) sample pig of the selection with advantage allele genotype carries out the selection and breeding of advantage strain.
6. application according to claim 5, it is characterised in that selection genotype carries out advantage product for pig homozygous G/G
The selection and breeding of system.
7. according to claim 5~6 any one of them application, it is characterised in that the advantage strain shows as lean meat percentage height.
8. according to claim 5~6 any one of them application, it is characterised in that the step (1) can use direct Sequencing,
Or first expand the mode that the fragment containing the SNP site detects again and carry out.
9. for expanding the primer pair containing SNP site fragment described in claim 1, it is characterised in that for from SEQ ID No.1
The fragment containing the SNP site is arrived in amplification in shown sequence.
10. claims 1 to 3 any one of them SNP site is in market pig/Native Pig economic characters advantage strain is identified
Using.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110117665A (en) * | 2019-05-15 | 2019-08-13 | 华南农业大学 | SNP marker relevant to pig lean meat percentage and eye muscle area and application on No. 16 chromosomes of pig |
| CN111709742A (en) * | 2020-05-18 | 2020-09-25 | 杜晓楠 | Method for preventing root address exposure in blockchain system, computer readable storage medium and blockchain system |
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| CN104480108A (en) * | 2014-12-12 | 2015-04-01 | 中国农业科学院北京畜牧兽医研究所 | Method for identifying pig back fat thickness and special primer pair thereof |
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| CN101824416A (en) * | 2010-04-30 | 2010-09-08 | 湖南农业大学 | Pork quality trait related gene DGAT1 and application thereof in porcine marker-assisted selection |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110117665A (en) * | 2019-05-15 | 2019-08-13 | 华南农业大学 | SNP marker relevant to pig lean meat percentage and eye muscle area and application on No. 16 chromosomes of pig |
| CN110117665B (en) * | 2019-05-15 | 2020-10-16 | 华南农业大学 | SNP molecular marker located on pig No. 16 chromosome and related to pig lean meat percentage and eye muscle area and application |
| CN111709742A (en) * | 2020-05-18 | 2020-09-25 | 杜晓楠 | Method for preventing root address exposure in blockchain system, computer readable storage medium and blockchain system |
| CN111709742B (en) * | 2020-05-18 | 2023-08-01 | 杜晓楠 | Method for preventing root address exposure in blockchain system, computer readable storage medium and blockchain system |
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