CN103667279A - Molecular marker of gene Resistin related to average daily gain of pigs and application of molecular marker - Google Patents

Abstract

The invention relates to the field of pig molecular marker preparation, in particular to a molecular marker of a gene Resistin related to average daily gain of pigs and application of the molecular marker. According to the invention, one SNP locus is contained in the Resistin gene sequence, is positioned the 861bpth position of the nucleotide sequence SEQ ID NO.1, is a basic group mutation of T/C and causes the Alw26I-RFLP polymorphism. The basic group mutation is positioned in the third intron of the Resistin gene without changing the translated amino acid sequence. A primer is designed in the vicinity of the SNP locus of the Resistin gene sequence to obtain the molecular marker sequence SEQ ID NO.1, so characters of the average daily gain of the pigs can be identified. The existing PCR (polymerase chain reaction)-RFLP (restricted fragment length polymorphisms) technology is used, one T-C mutation is found at the 861bpth position of the Resistin gene, the polymorphic site is found to be related to the characters of the average daily gain and a certain help is provided for the research on the pig growth characters.

Description

Molecule marker and the application thereof of pig average daily gain genes involved Resistin

Technical field

The present invention relates to pig molecule mark preparation field, refer to particularly molecule marker and the application thereof of a boar average daily gain genes involved Resistin.

Background technology

Livestock industry is in occupation of very important status in modern agricultural development, and its ratio occupying in agriculture production is commonly used to weigh the important indicator of a countries and regions development degree.Pork is as the topmost meat of China source, pig-breeding industry in China's livestock industry in occupation of very important status.At 20th century the mid-80, approximately 2,000 ten thousand tons of China's productions of meat, pork accounts for 85%, 2012 year pork output and has reached 5,335 ten thousand tons, accounts for meat total amount 63.63%.Although pork proportion declines gradually, it is dominate all the time.

In the industry of raising pigs, feed cost accounts for the 65-75% of the total cost of raising pigs.Therefore, reduce the feed intake in pig-breeding, improve food utilization efficiency, reduce feed cost and be related to raise pigs enterprise development and even existence.Research discovery, and the economic characters significant correlations such as the same food consumption of feed efficiency of pig, average daily gain, intramuscular fat (Zhao Kebin improves the strategy that growing-finishing pig feed conversion rate reduces feed cost. pig industry science, and 2008,60-63).Therefore the gene that, affects the proterties such as pig feed intake, average daily gain, intramuscular fat can be used as the candidate gene of research pig feed transformation efficiency.

Phylaxin Resistin is by (Steppan CM such as the Steppan of Univ Pennsylvania USA, Bailey ST, Bhat S, Brown EJ, Banerjee RR, Wright CM, Patel HR, Ahima RS, Lazar MA. " The hormone resistin links obesity to diabetes " [J] .Nature, 2001,409 (6818): 307 – 312) in ripe adipocyte, find.The same year, Kim etc. utilize the separation of gene chip isolation technique to obtain Resistin gene (the Sung-Eun Kim of mouse, He Huang, Ming Zhao, Xinjun Zhang, Aili Zhang.Wnt Stabilization of β-Catenin Reveals Principles for Morphogen Receptor-Scaffold Assemblies[J] .Science, Published Online April112013).Subsequently, the poor screening library separating out fat cell that subtracts of the utilizations such as Rajala is when the gene of different differential periods expression, also isolated Resistin gene (Rajala MW, Lin Y, Ranalletta M, Yanq XM, Qian H, Ginqerich R.Cell type-specific expression and coregulation of murine resistin and resistin-like molecule-alpha in adipose tissue[J] .Mol Endocrinol, 2002,16:1920 – 30).In the promotor of the employing PCR-SSCP method discovery Resistin genes such as Pan Yingbin, exon, intron, all find polymorphic site (Pan Yingbin, Liu Yanfen, Liu You, Gao Zhijie, Wang Qun, He Wanjuan, Tan Minjing. six kind pork insulin resistin gene single nucleotide polymorphism analysis. Journal of Agricultural Biotechnology, 2008,16 (3): 402-406).

Model red flag by the research of insulin sensitivity target tissue-Skeletal Muscle Cell Sugar intake function is shown Resistin can significantly suppress myocyte on glucose uptake (model red flag .Resistin is on the impact of Skeletal Muscle Cell Sugar intake function and mechanism analysis .[master thesis thereof], Nanjing: Nanjing Medical University, 2007).The researchs such as Solana show mouse Resistin gene can with receptor tyrosine kinase sample orphan receptor 1(ROR1) in specific region, ROR1 extracellular, be combined, mediation 3T3-L1 preadipocyte differentiation and glucose uptake (Beatriz S á nchez-Solana, Jorge Laborda, Victoriano Baladr ó n.Mouse Resistin Modulates Adipogenesis and Glucose Uptake in3T3-L1Preadipocytes Through the ROR1Receptor[J] .Molecular Endocrinology, 2012,26 (1): 110-127).Liu Feng etc. found by further research that expression Resistin slowed down beta Cell of islet RINm5F and breeds, show that Resistin can produce restraining effect (Liu Feng to insulin secretion, Qiu Jie, Zhang Chunmei, Ji Chenbo, the impact of the molten .Resistin gene pairs beta Cell of islet RINm5F propagation of Guo's tin. Chinese Journal of Contemporary Pediatrics, 2010,43-46).

Resistin can be used as signaling molecule and regulates fat growth growth and the deposition that distributes simultaneously.Chen little Dong detects the relevant parameter of some steatolysis in Food restriction and the pork fat of being satiated with food, in the fatty tissue of discovery pig, Resistin significantly expresses, and Resistin expression level with fatty deposits be proportionate (Chen little Dong. the expression of Adipocyte Factor TNF-α and resistin and the regulating and controlling effect .[doctorate paper to pork fat metabolism thereof], Wuhan: Hua Zhong Agriculture University, 2004).Cieslak finds in Polish Large White colony, Resistin gene is with thickness of backfat significant correlation (Cieslak J, Nowacka-Woszuk J, Bartz M, Fijak-Nowak H, Grzes M, Szydlowski M, Switonski M.Association studies on the porcine RETN, UCP1, UCP3and ADRB3genes polymorphism with fatness traits[J] .Meat Science, 2009,83 (3): 551-554).

More than research shows, Resistin can be by insulin action, and blood-sugar content in control agent, simultaneously with propagation, the differentiation of body fat cell and have close ties with fatty deposition level, by regulating Resistin expression amount, can control volume self-energy metabolic balance.

Because carbohydrate metabolism and fatty deposits in the growth traits of pig and body have substantial connection, therefore can study Resistin gene and whether pig growth traits be exerted an influence.Because Resistin is regulating the important role (Yang Qing such as lipid metabolism, insulin resistance, energy balance, food consumption, inflammation, Zhou Lei, Gan Li, thunderclap, expression and the study on regulation of old little winter .Resistin gene. biochemical the tenth the seminar abstract of a thesis compilation of national animal physiological, 2008), research Resistin and the energy metabolism of pig, the relation between food consumption, significant for the economic characters of improvement pig.

Summary of the invention

Technical problem to be solved by this invention is just to provide the molecule marker of a kind of pig average daily gain genes involved Resistin, and its nucleotide sequence is as shown in SEQ ID NO.2.

It is a kind of for obtaining the primer pair of described molecule marker that the present invention also provides, and described primer pair is:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 '.

The present invention also provides a kind of preparation method of molecule marker, and the pig genomic dna with Resistin gene of take is template, adopts following primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 '.

Carry out pcr amplification, its molecule marker obtaining is SEQ ID NO.2.

The present invention also provides the method for the high average daily gain proterties of a kind of molecular markers for identification pig, and the genomic dna of pig to be measured of take is template, adopts following primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

Carry out pcr amplification, wherein, have the pig of 349bp band, it has Resistin gene, as SEQ ID NO.1.

The present invention also provides the application of a kind of molecule marker in pig some growth proterties and feed efficiency proterties related molecular marker assistant breeding.

The application of the primer pair that the present invention also provides a kind of molecule marker in pig some growth proterties and feed efficiency proterties related molecular marker assistant breeding.

A method for the pig that evaluation contains Resistin gene, comprises the following steps and is:

1) primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

2) pcr amplification condition

PCR reaction cumulative volume 10 μ l, pig genomic dna template 1 μ l to be measured wherein, each 0.2nmol/ μ l of forward and reverse primer, PCRmix5 μ l, finally adds deionized water to cumulative volume 10 μ l;

3) PCR reaction conditions is: after 95 ℃ of denaturation 5min, 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s circulate; Last 72 ℃ are extended 5min; PCR product detects through 2%W/V agarose gel electrophoresis;

4) RFLP detects

PCR product endonuclease reaction volume is 10 μ l, PCR product 5 μ l wherein, and deionized water 3.5 μ l, 10 * buffer1 μ l, restriction enzyme A lw26I is 0.5 μ l (10U/ μ l),

By centrifugal after sample blending, 37 ℃ of incubators are placed 12h, detect enzyme cut result with 3.5%W/V agarose gel electrophoresis, record genotype, under ultraviolet lamp, take pictures;

Enzyme is cut and is produced three kinds of genotype, and TT genotype only has 349bp mono-band, and CC genotype has 183bp and 166bp two bands, and heterozygote TC genotype has 349bp, 183bp and 166bp tri-bands.

The principle of the invention

In Resistin gene order, comprise a SNP site, be positioned at the 861bp place of nucleotide sequence SEQ IDNO.1, the base mutation that this site is T/C, causes Alw26I-RFLP polymorphism.This base mutation is arranged in Resistin gene the 3rd intron, does not change the aminoacid sequence of its translation.Therefore design primer near the SNP site of Resistin gene order, obtain molecule labelled series SEQ ID NO.1, thereby identify pig average daily gain proterties.

In order to realize above-mentioned object, the present invention adopts following technical measures:

A kind of acquisition of the molecule marker relevant to pig some growth proterties and feed efficiency proterties:

1) design of primers of pig Resistin gene and partial dna sequence amplification

With pig Resistin gene order information (the GenBank number of including: ENSSSCG00000013575) as the template sequence of design of primers, utilize biology primer-design software Oligo7.0 design primer, primer sequence is as follows:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

2) pcr amplification reaction:

Utilizing TIANamp Genomic DNA Kit test kit (purchased from U.S. invitrogen company) is that (this colony derives from Hubei gold woods breeding station to great Bai castrating pig experimental population from purebred U.S.A,) (slaughter determining and sampling are divided into 21 batches and carry out, measure individual amount and amount to 236, be all that U.S.A is purebred great Bai castrating boar) extract genomic dna in ear tissue, concrete operation method carries out with reference to TIANamp Genomic DNA Kit test kit specification sheets.

PCR reaction: reaction cumulative volume is 30 μ l, pig DNA template 3.0 μ l wherein, PCRmix15 μ l, each 0.6nmol/ μ l of forward primer and reverse primer, finally adds deionized water to cumulative volume 30 μ l.PCR reaction conditions is: after 95 ℃ of denaturation 5min, and circulate 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s, last 72 ℃ are extended 5min.PCR product detects through 1% (W/V) agarose gel electrophoresis, has obtained part Resistin fragment, and length is 349bp, and its sequence is shown in SEQ ID NO.2.

3) purifying of PCR product, order-checking

The purifying of PCR product: cut the gel containing object fragment from low melting-point agarose gel under ultraviolet lamp, put into 1.5ml centrifuge tube, then use PCR product purification test kit (purchased from Beijing hundred Imtech) purified pcr product.DNA solution after purifying is delivered to the forward and reverse order-checking of Shanghai Ying Jun company.

With primer amplification pig genomic dna, obtained 349bp specific amplification fragment, as shown in Figure 2, forward and reverse sequencing result is found a 861st T-C conversion of discovery (as shown in Figure 3) of the Resistin gene (shown in SEQ ID NO.1) at this fragment place, causes an Alw26I restriction enzyme site .

A detection method for molecule marker based on relevant to pig average daily gain, its step is as follows:

1) primer sequence

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

This primer amplification fragment length 349bp, its sequence is shown in SEQ ID NO.2.

2) pcr amplification condition

PCR reaction cumulative volume 10 μ l, pig genomic dna template 1 μ l wherein, each 0.2nmol/ μ l of forward and reverse primer, PCRmix5 μ l, finally adds deionized water to cumulative volume 10 μ l.PCR reaction conditions is: after 95 ℃ of denaturation 5min, 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s circulate; Last 72 ℃ are extended 5min.PCR product is through 2%(W/V) agarose gel electrophoresis detection.

3) RFLP detects

PCR product endonuclease reaction volume is 10 μ l, PCR product 5 μ l wherein, deionized water 3.5 μ l, 10 * buffer1 μ l, restriction enzyme A lw26I is 0.5 μ l (10U/ μ l), by centrifugal after sample blending, 37 ℃ of incubators are placed 12h, with 3.5%(W/V) agarose gel electrophoresis detects enzyme and cuts result, records genotype, under ultraviolet lamp, take pictures.

Enzyme is cut and is produced three kinds of genotype, and TT genotype only has 349bp mono-band, and CC genotype has 183bp and 166bp two bands, and heterozygote TC genotype has 349bp, 183bp and 166bp tri-bands.

To the molecule marker of pig average daily gain proterties with pig some growth proterties and the relevant molecular mark of feed efficiency proterties in an application, step is as follows:

The average daily gain of collected specimens, food consumption, prediction food consumption, residue food consumption, feed conversion rate and 6 indexs of intramuscular fat.According to the group structure of collected specimens, use mixture model come statistical study Resistin gene SNP site genotype effect and with the relation of feed efficiency proterties:

Y _ij=μ+genotypei+ε _ij+G

Wherein, Y _ijfor processing rear character value, μ is population mean, and ε ij is random error, and G is a batch effect, supposes and obeys N(0, σ ²) distribute.Can analyze genotype effect, complete the multiplicity comparison of genotype effect.Adopt SPSS16.0 software (AsiaAnalytics China) to carry out data processing and statistical study.

The T>C mutator gene type of this gene is found in statistical study, and wherein TT type and the different of CC type and average daily gain significant correlation (P=0.038), with food consumption, prediction food consumption and residue food consumption significant correlation not.The individuality that is TT, CC, TC to genotype by least squares means compares between two, and result shows that TT type genotypic mean day weight gain is individual higher than CC type genotype, between TC type genotype and other two kinds of genotype without significant difference.

Compared with prior art, the present invention has the following advantages:

The present invention utilizes existing PCR-RFLP technology, finds a T-C sudden change, and find this polymorphic site and average daily gain proterties significant correlation at Resistin gene 861bp place, and the research of pig growth traits is had to certain help.

Accompanying drawing explanation

Fig. 1 is the technology of the present invention schema.

Fig. 2 is pig Resistin gene genome amplification electrophoresis result schematic diagram.

M:DL2000 molecular weight marker;

Fig. 3 is the T>C sudden change that in the present invention, pig Resistin gene sequencing is found.

Fig. 4 is three kinds of genotype (TT CC TC) electrophoresis result schematic diagram of the Alw26I-RFLP of pig Resistin gene extron.

M:50bpDNA molecular weight marker; P:PCR product.

Embodiment

In order to explain better the present invention, below in conjunction with specific embodiment, further illustrate main contents of the present invention, but content of the present invention is not only confined to following examples.

Embodiment 1:

A kind of acquisition of the molecule marker relevant to pig average daily gain proterties:

(1) design of primers of pig Resistin gene and partial dna sequence amplification

With pig Resistin gene order information (the GenBank number of including: ENSSSCG00000013575) as the template sequence of design of primers, utilize biology primer-design software Oligo7.0 design primer, primer sequence is as follows:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

(2) pcr amplification reaction:

Utilizing TIANamp Genomic DNA Kit test kit (purchased from U.S. invitrogen company) is that (slaughter determining and sampling are divided into 21 batches and carry out great Bai castrating pig experimental population (this colony derives from Hubei gold woods breeding station) from purebred U.S.A, measure individual amount and amount to 236, be all that U.S.A is purebred great Bai castrating boar) extract genomic dna in ear tissue, concrete operation method carries out with reference to TIANamp Genomic DNA Kit test kit specification sheets.

PCR reaction: reaction cumulative volume is 30 μ l, pig DNA template 3.0 μ l wherein, PCRmix15 μ l, each 0.6nmol/ μ l of forward primer and reverse primer, adds deionized water to cumulative volume 30 μ l.PCR reaction conditions is: after 95 ℃ of denaturation 5min, and circulate 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s, last 72 ℃ are extended 5min.PCR product is through 1(W/V) detection of % agarose gel electrophoresis, obtained part Resistin fragment, length is 349bp, its sequence is shown in SEQ ID NO.2.

(3) purifying of PCR product, order-checking

The purifying of PCR product: cut the gel containing object fragment from low melting-point agarose gel under ultraviolet lamp, put into 1.5ml centrifuge tube, then use PCR product purification test kit (purchased from Beijing hundred Imtech) purified pcr product, according to test kit specification sheets, operate, concrete steps are to add the ratio of 400 μ l GS Buffer to add GS Buffer according to 100mg blob of viscose, put 50 ℃ of incubation 10min, agarose blob of viscose is dissolved completely, within every two minutes, put upside down and mix once; The glue dissolving is transferred to centrifugal adsorbing column, and centrifugal adsorbing column is placed in to waste collection pipe, the centrifugal 30s of 10000rpm, discards waste liquid; Centrifugal adsorbing column is put back in waste collection pipe, added 500 μ l Wash Buffer in centrifugal adsorbing column, the centrifugal 30s of 10000rpm, abandons filtrate.With same method, use again 500 μ l Wash Buffer solution washings once; Centrifugal adsorbing column is put in meeting waste collection pipe to the centrifugal 1min of top speed; The careful centrifugal adsorbing column that takes out, is inserted in an aseptic 1.5ml centrifuge tube, in adsorption film central authorities, adds 30 μ l distilled waters, after the standing 2-10min of room temperature, and the centrifugal 1min of top speed; Take out centrifugal adsorbing column, 1.5ml centrifuge tube (DNA solution) is placed in to-20 ℃ and saves backup.DNA solution after purifying is delivered to the forward and reverse order-checking of Shanghai Ying Jun company.

With primer amplification pig genomic dna, obtained 349bp specific amplification fragment, as shown in Figure 2, forward and reverse sequencing result is found a 861st T-C conversion of discovery (as shown in Figure 3) of the Resistin gene (shown in SEQ ID NO.1) at this fragment place, causes an Alw26I restriction enzyme site

Embodiment 2:

A detection method for molecule marker based on relevant to pig average daily gain proterties, its step is as follows:

PCR-RFLP diagnostic method is set up

(1) primer sequence

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

This primer amplification fragment length 349bp, its sequence is shown in SEQ ID NO.2.

(2) pcr amplification condition

PCR reaction cumulative volume 10 μ l, pig genomic dna template 1 μ l wherein, each 0.2nmol/ μ l of forward and reverse primer, PCRmix5 μ l, finally adds deionized water to cumulative volume 10 μ l.PCR reaction conditions is: after 95 ℃ of denaturation 5min, 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s circulate; Last 72 ℃ are extended 5min.PCR product is through 2%(W/V) agarose gel electrophoresis detection.

(3) RFLP detects

PCR product endonuclease reaction volume is 10 μ l, PCR product 5 μ l wherein, deionized water 3.5 μ l, 10 * buffer1 μ l, restriction enzyme A lw26I is 0.5 μ l (10U/ μ l), by centrifugal after sample blending, 37 ℃ of incubators are placed 12h, with 3.5%(W/V) agarose gel electrophoresis detects enzyme and cuts result, records genotype, under ultraviolet lamp, take pictures.

Enzyme is cut and is produced three kinds of genotype, and TT genotype only has 349bp mono-band, and CC genotype has 183bp and 166bp two bands, and heterozygote TC genotype has 349bp, 183bp and 166bp tri-bands, as shown in Figure 4.

Embodiment 3:

The application of the molecule marker relevant from pig average daily gain proterties in different swinery body polymorphic detections, the steps include:

Utilizing PCR-Alw26I-RFLP to detect purebred U.S.A is great Bai castrating pig experimental population (this colony derives from Hubei gold woods breeding station) (236 U.S.As are that purebred great Bai castrates boar colony).Genotype and the gene frequency of this mutational site in experimental population is as shown in table 1, detected result shows, in experimental population, there are three kinds of genotype in Resistin gene, wherein TT type individuality has 104, TC type individuality has 112, CC type individuality has 20, and the detected result of enzyme cutting type conforms to sequencing result, and detection method of the present invention is reliable.This result shows that Resistin gene is that in great Bai castrating pig experimental population, allelotrope T preponderates in purebred U.S.A.

Embodiment 4:

The association analysis of a kind of molecule marker relevant to pig average daily gain proterties and some growth proterties and feed efficiency proterties

The swinery of the present embodiment is great Bai castrating pig from this Hubei purebred U.S.A of gold woods breeding station, and slaughter determining and sampling are divided into 21 batches to be carried out, and measures individual amount and amounts to 236, is all that U.S.A is purebred great Bai castrating boar.

The proterties of analyzing is mainly some growth proterties and the proterties relevant to feed efficiency proterties, comprising: average daily gain, food consumption, prediction food consumption, residue food consumption, feed conversion rate and 6 indexs of intramuscular fat.According to the group structure of collected specimens, applicant use mixture model come statistical study Resistin gene SNP site genotype effect and with the relation of some growth and feed efficiency proterties:

Yij=μ+genotypei+ε _ij+G

Wherein, Yij is character value after processing, and μ is population mean, and ε ij is random error, and G is a batch effect, supposes and obeys N(0, σ ²) distribute.Can analyze genotype effect, complete the multiplicity comparison of genotype effect.Adopt SPSS16.0 software (software comes from AsiaAnalytics China) to carry out data processing and statistical study.

Pig Resistin Gene A lw26I-RFLP pleomorphism site and some growth and feed efficiency proterties are carried out to association analysis, known: TT type individuality has 104 in 236 individualities by table 1, TC type individuality has 112, and CC type individuality has 20.

It is, in great Bai castrating swinery body, some growth and feed efficiency proterties have been carried out to preliminary association analysis to the polymorphic of the T>C mutational site of pig Resistin gene in U.S.A that the present embodiment adopts the generalized linear model in SPSS16.0 software.Preliminary Analysis Results is in Table 1.The T>C mutator gene type of this gene is found in statistical study, and wherein TT type and the different of CC type and average daily gain significant correlation (P=0.038), with food consumption, prediction food consumption and residue food consumption significant correlation not.The individuality that is TT, CC, TC to genotype by least squares means compares between two, and result shows that TT type genotypic mean day weight gain is individual higher than CC type genotype, between TC type genotype and other two kinds of genotype without significant difference.

The association analysis of table 1Resistin gene polymorphism sites genotype and some growth proterties and feed efficiency proterties detects

Note: the proterties average in table is mean number ± standard deviation.

ADG: average daily gain; FI: daily ingestion amount; PFI: prediction daily ingestion amount; RFI: residue food consumption; FCR: feed conversion rate; IMF: intramuscular fat.

Other unspecified part is prior art.Although above-described embodiment has been made detailed description to the present invention; but it is only the present invention's part embodiment; rather than whole embodiment, people can also obtain other embodiment according to the present embodiment under without creative prerequisite, and these embodiment belong to protection domain of the present invention.

Claims (7)

1. the molecule marker of a boar average daily gain genes involved Resistin, is characterized in that: its nucleotide sequence is as shown in SEQ ID NO.2.

2. for obtaining the primer pair of molecule marker described in claim 1, it is characterized in that: described primer pair is:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 '.

3. by the preparation method of the molecule marker described in claim 1 or 2, it is characterized in that: the pig genomic dna with Resistin gene of take is template, adopts following primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 '.

Carry out pcr amplification, its molecule marker obtaining is SEQ ID NO.2.

4. according to the method for the high average daily gain proterties of molecular markers for identification pig claimed in claim 1, it is characterized in that: the genomic dna of pig to be measured of take is template, adopts following primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

Carry out pcr amplification, wherein, have the pig of 349bp band, it has Resistin gene, as SEQ ID NO.1.

5. the application of molecule marker claimed in claim 1 in pig some growth proterties and feed efficiency proterties related molecular marker assistant breeding.

6. the application of the primer pair of molecule marker claimed in claim 2 in pig some growth proterties and feed efficiency proterties related molecular marker assistant breeding.

7. a method for the pig that evaluation contains Resistin gene, is characterized in that, comprises the following steps to be:

1) primer pair:

Forward primer: 5 '-CCCTCAGGTGAGTACAGAACT-3 ';

Reverse primer: 5 '-AAGCCTGCAAGTGGGAATGAG-3 ';

2) pcr amplification condition

PCR reaction cumulative volume 10 μ l, pig genomic dna template 1 μ l to be measured wherein, each 0.2nmol/ μ l of forward and reverse primer, PCRmix5 μ l, finally adds deionized water to cumulative volume 10 μ l;

3) PCR reaction conditions is: after 95 ℃ of denaturation 5min, 30 95 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ of extension 40s circulate; Last 72 ℃ are extended 5min; PCR product detects through 2%W/V agarose gel electrophoresis;

4) RFLP detects

PCR product endonuclease reaction volume is 10 μ l, PCR product 5 μ l wherein, and deionized water 3.5 μ l, 10 * buffer1 μ l, restriction enzyme A lw26I is 0.5 μ l (10U/ μ l),

By centrifugal after sample blending, 37 ℃ of incubators are placed 12h, detect enzyme cut result with 3.5%W/V agarose gel electrophoresis, record genotype, under ultraviolet lamp, take pictures;

Enzyme is cut and is produced three kinds of genotype, and TT genotype only has 349bp mono-band, and CC genotype has 183bp and 166bp two bands, and heterozygote TC genotype has 349bp, 183bp and 166bp tri-bands.

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