CN113308549A - Molecular marker of chicken feather color trait related gene MC1R and application thereof - Google Patents

Molecular marker of chicken feather color trait related gene MC1R and application thereof Download PDF

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CN113308549A
CN113308549A CN202110541670.4A CN202110541670A CN113308549A CN 113308549 A CN113308549 A CN 113308549A CN 202110541670 A CN202110541670 A CN 202110541670A CN 113308549 A CN113308549 A CN 113308549A
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王艳
吴日富
瞿浩
舒鼎铭
刘天飞
罗成龙
计坚
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Institute of Animal Science of Guangdong Academy of Agricultural Sciences
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Abstract

The invention discloses a molecular marker of chicken feather color trait related gene MC1R and application thereof. Relates to the technical field of chicken marker-assisted selection. Compared with the MC1R gene coding region DNA sequence, the sequence comprises the following components: SNP 1-SNP 5. The method selects the feather color characters of different types of black feathers and black feathers, accelerates the feather color selection breeding process and improves the feather color consistency of chicken flocks, and when SNP 1-SNP 5 is CTGAC, the proportions of the black feather type of the chest and the black-feathered phenotype of the back of an F2-generation cock are improved; when SNP 1-SNP 5 is CTGAG, the proportion of yellow feather type at the chest part, yellow x jute feather type at the back part of the F2 hen is improved; when SNP 1-SNP 5 is TCACC, the proportions of the breast tingling and back tingling and feathering phenotype of the F2 generation hen and the breast red feather and back red and feathering phenotype of the F2 generation cock are improved.

Description

Molecular marker of chicken feather color trait related gene MC1R and application thereof
Technical Field
The invention relates to the technical field of chicken marker-assisted selection, in particular to a molecular marker of a chicken feather color trait related gene MC1R and application thereof.
Background
Local chicken varieties in China are rich in resources, are deeply loved by consumers due to delicious meat quality and have great potential as breeding materials, but due to the fact that the feather colors in the varieties are different, factors such as genetic rules and mechanism research on the feather colors are not deep and the like can greatly influence the utilization of good varieties and the economic benefit of commodities.
For a long time, the feather color of birds and the hair color and skin color of mammals are the key points of pigment research. Feather color can be used as a genetic marker, and is helpful for identifying varieties, populations and breeding populations according to characteristics of the feather. The modern molecular technology is used for screening the molecular marker of the candidate gene of the feather color of the poultry, so that the breeding process and the uniformity of the feather color can be improved, the breeding time is effectively saved, and the economic benefit is improved. The feather-colored pigment of chicken is mainly formed by melanin and carotenoid, wherein the carotenoid needs to be taken from the outside, and the melanin can be synthesized by itself. Melanin is further divided into eumelanin, which makes the feathers black, and pheomelanin, which makes the feathers reddish brown and yellow, and pheomelanin, both of which are tyrosine derivatives; generally, the color of the feather of chicken is mainly formed by the difference of the amount, proportion and position of eumelanin and pheomelanin in the feather.
The color of chicken feathers is regulated by a number of genes, some of which cause the primary effect of color, and others of which act to modify and regulate, including affecting pigment content and proportion and distribution in individual feathers. Among the most studied are eumelanin expansion locus E (E × tension, E) which encodes the melanocortin receptor MC1R, which regulates the relative proportion and distribution of eumelanin and pheomelanin, and MC1R which influences the synthesis of eumelanin and pheomelanin by regulating the activity of tyrosine.
However, at present, from the genetic point of view, a detection method related to the MC1R gene as a chicken feather color character molecular marker is not established, and the gene as the chicken feather color character molecular marker for auxiliary selection is not applied.
Therefore, the problem to be solved by those skilled in the art is how to provide a molecular marker of gene MC1R related to chicken feather color trait.
Disclosure of Invention
In view of the above, the invention provides a molecular marker of chicken feather color trait related gene MC1R and application thereof. The invention provides reference for evaluation of the feather color character in the germ plasm resource depth and breeding utilization. The molecular marker is located on the coding region of the chicken MC1R gene. Specifically, the molecular marker is a marker located in SEQ ID NO: 1, the C/T base mutation at 69 th base, the T/C base mutation at 212 th base, the G/A base mutation at 274 th base, the A/C base mutation at 644 th base and the C/G base mutation at 919 th base in the sequence.
In order to achieve the purpose, the invention adopts the following technical scheme:
a molecular marker of chicken feather trait related gene MC1R, compared with MC1R gene coding region DNA sequence SEQ ID NO.1, comprises: SNP1 at position 69, SNP2 at position 212, SNP3 at position 274, SNP4 at position 644 and SNP5 at position 919;
when the SNP1 is C, SNP2, T, SNP3, G, SNP4 and A, SNP5 are C, the chicken is haplotype H1, and the proportion of the black feather type of the chest and the black-tinged feather type of the back of the F2 cock is improved;
when the SNP1 is C, SNP2, T, SNP3, G, SNP4 and A, SNP5 are G, the chicken is haplotype H2, and the proportion of the yellow feather type of the breast part, yellow of the back part and the yellow feather type of the jute of the F2 generation hen is improved;
when the SNP1 is T, SNP2, C, SNP3, A, SNP4 and C, SNP5 which are C, the chicken is haplotype H3, and the proportions of the F2 generation hen chest pockmarked feather, back pockmarked feather and F2 generation cock chest red feather, back pockmarked feather and C are improved.
Preferably: the primers used for amplifying the MC1R gene coding region DNA sequence SEQ ID NO. 1:
a forward primer MC1R-2F: 5'-CTTCCCCATCCTTGTGCCTG-3' as shown in SEQ ID NO. 2;
a reverse primer MC1R-2R: 5'-CCTTTATTTGGGAGCGCGAG-3' as shown in SEQ ID NO. 3;
the invention also provides application of the molecular marker in preparation of a kit for detecting the feather color of chicken.
The invention also provides application of the molecular marker in chicken breeding.
The invention also provides a method for detecting the molecular marker of the gene MC1R related to the feather color character of the chicken, which comprises the following steps:
(1) performing PCR amplification by using chicken blood DNA as a template;
(2) and carrying out SNP screening on the amplified product to construct a haplotype.
Preferably: step (1) PCR amplification system: in a PCR reaction system with a total volume of 25. mu.L, 1. mu.L of a chicken blood DNA template, 12.5. mu.L of a 2 XPCR reaction mixture, 1. mu.L of each of 10mM forward and reverse primers, 0.25. mu.L of Taq DNA polymerase and 9.25. mu.L double distilled water were added.
Preferably: the reaction conditions of the PCR in the step (1) are as follows: pre-denaturation at 94 ℃ for 4 min; denaturation at 94 ℃ for 30s, annealing at 62 ℃ for 30s, and extension at 72 ℃ for 1min20s for 34 cycles; finally, the extension is carried out for 10min at 72 ℃ and the product is stored at 4 ℃.
According to the technical scheme, compared with the prior art, the molecular marker of the gene MC1R related to the chicken feather color character and the application thereof are disclosed, and the technical effects that the molecular marker can be used for selecting different types of black feather and pockmarked feather color characters, accelerating the feather color selection breeding process and improving the consistency of the feather color of chicken flocks are achieved.
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In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the provided drawings without creative efforts.
FIG. 1 is a diagram showing the DNA sequence of the coding region of the chicken MC1R gene in example 1 of the present invention, and an electrophoretogram (agarose gel concentration of 1.5%) of a fragment for searching for a molecular marker, wherein M: DS2000 DNA MAKER, lanes 1-5: and (4) amplifying the product.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The embodiment of the invention discloses a molecular marker of a chicken feather color trait related gene MC1R and application thereof.
Example 1
Establishment of molecular detection method for chicken black feather and pockmarked feather color characters
(one) primer:
CCCGTGCAAGCGCCTGCGCCTGCAAGGGCCCGTCCAGGCCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGGCCATCGGCCCTGGCCCTGCGTCCGTCCGTCCGGCCCTGCAAGGGCCCGTCCGTGCCTGCAGGCCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGGCCCTGCAAGGGCCTGCAGGCCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGTCCGGCCCTGCGGCCCTGCGGCCCTGCGGCCGTCCGTGCGGCCCTGCGTGCGGCCCCGTGCGGCCCTGGCCCTGCGTGCGTCCGGCCCTGCGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCGGGCCCGGCCCCGGCCCTGGCCGGCCCTGGCCGTCCGGCCGGCCGTCCGTCCGGCCCTGGCCGTCCGTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGGCCCTGCCTGCCTGGCCCTGCCTGGCCCTGGCCCTGCCTGGCCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGGCGGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGGCCCTGGCCCTGGCCCTGCCTGCCTGCCTGCCTGGCCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCCTGGCCCTGGCCCTGCCTGGCCGGGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCGGGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCGGGCCTGGCCCTGGCCCTGGCCCTGCCTGGCCCTGGCCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCCTGCCTGCCTGGCCCTGGCCCTGGCCGGGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGGCCCTGCCTGCCTGGCCGGGCCTGCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGGCCCTGGCCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGGCCCTGGCCCTGCCTGCCTGGCCGGGCCTGGCCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGGCCCTGGCCCTGCCTGCCTGGCCCTGCCTGGCCCTGGCCCTGCCTGGCCCTGGCCCTGCCTGGCCCTGCGGGCCTGCGGGCCTGCGGGCCTGCCTGCCTGGCCCTGCCTGCCTGGCCGGGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGGCCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGGCCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGTCCGTCCGTCCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCGGGCCTGCCTGCGGGCCTGCCTGCCTGCGGGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTGCCTG. The sequences of the primers are respectively shown as SEQ ID NO.2 and SEQ ID NO.3, and the sequences are as follows:
a forward primer MC1R-2F: 5'-CTTCCCCATCCTTGTGCCTG-3' as shown in SEQ ID NO. 2;
reverse primer MC1R-2R: 5'-CCTTTATTTGGGAGCGCGAG-3' as shown in SEQ ID NO. 3.
(II) PCR amplification conditions:
to a PCR reaction system with a total volume of 25. mu.L, 1. mu.L of DNA template, 12.5. mu.L of 2 XPCR reaction mixture, 1. mu.L of each of 10mM forward and reverse primers, 0.25. mu.L of Taq DNA polymerase, and 9.25. mu.L of double distilled water were added, and PCR reaction conditions were as follows: pre-denaturation at 94 ℃ for 4 min; denaturation at 94 ℃ for 30s, annealing at 62 ℃ for 30s, and extension at 72 ℃ for 1min20s for 34 cycles; finally, the extension is carried out for 10min at 72 ℃ and the product is stored at 4 ℃.
(III) searching molecular markers:
directly sequencing the PCR amplification product, analyzing the sequence by using DNMAN software to find the base mutation at 69 th, 212 th, 274 th, 644 th and 919 th positions, constructing haplotypes for the 5 SNPs by using phase2.1 software, and performing association analysis on the haplotypes and the feather color characters by using a Haplo Stats software package in an R language environment.
(IV) Experimental materials:
the special strain R with the feather color of black chest and red back is used as a male parent, the special strain S with the brown feather character is used as a female parent, hybridization is carried out to generate F1 generation, and the F1 generation female parent is backcrossed with the R cock to generate a feather color separation backcross F2 population. 118 individuals of F2 adult cocks and 123 individuals of adult hens are randomly selected, classified according to the sex, the color characters of the chest and the back feathers, and the feather color of the chest and the back of each chicken is recorded in detail. 2ml of whole blood is collected through the subpteran vein by using an EDTA anticoagulation vacuum blood collection tube and stored in a refrigerator at the temperature of 20 ℃ below zero. The experimental materials are shown in table 1.
TABLE 1 Experimental materials information Table
Figure BDA0003072045810000051
Figure BDA0003072045810000061
(V) experimental procedures and results:
the experimental group blood DNA shown in Table 1 was selected as a template for PCR amplification, and the PCR product was detected by 1.5% agarose gel electrophoresis, with the detection results shown in FIG. 1. FIG. 1 shows the partial chicken MC1R gene coding region sequence as SEQ ID NO: 1 (M is DS2000Maker, and 1-5 is a target sequence band 1308bp of a partial sample). The PCR amplification product is sent to the forward and reverse sequencing of the company Limited in the biological engineering (Shanghai) to splice the sequencing result.
SNP screening is carried out on a sequence obtained by sequencing by using DNAMAN software, and 5 SNPs are respectively shown as SEQ ID NO: 1 (C69T), 212 (T212C), 274 (G274A), 644 (A644C) and 919 (C919G).
Haplotype was constructed for these 5 SNPs using phase2.1 software, and the results are shown in table 2, and haplotype frequency analysis was performed for 4 different populations, and the results are shown in table 3. And (3) performing correlation analysis on haplotypes and different feather color traits obtained by analyzing the phase2.1 software by utilizing a Haplo Stats software package in an R language environment. The software detects the relevance of the haplotypes and the traits by using a generalized linear model, the returned P value represents the significance of the relevance of the traits and the haplotypes, the score of the haplotype effect and the positive and negative of the haplotype effect represent the magnitude and the positive and negative of the haplotype effect, the score is positive, the positive value represents that the positive sequence trait effect of the feather color trait score sorting is large (namely the trait relevance corresponding to the highest feather color trait sorting score is higher), the score is negative, and the larger the absolute value is, the reverse sequence trait effect of the feather color trait score sorting is large (namely the trait relevance corresponding to the lowest feather color trait sorting score is higher); the P value of 0.05 or less is significant, and the P value of 0.01 or less is extremely significant.
In the experiment, according to the feather color of the breast, the back and the breast-back combined phenotype, the feather color characters of the F2 cock and the hen are respectively sequenced from deep to light, and then are associated with the haplotype for analysis. The results of the ranking of the different part feather phenotype are shown in Table 4, and the results of the correlation analysis between the different part feather phenotype and the haplotype are shown in Table 5.
TABLE 2 haplotype results of 5 SNPs on MC1R gene
Figure BDA0003072045810000071
TABLE 3 distribution of MC1R Gene haplotypes in different lines
Figure BDA0003072045810000072
TABLE 4 results of depth-to-light ordering of the feather phenotype at different sites
Figure BDA0003072045810000073
Figure BDA0003072045810000081
TABLE 5 correlation analysis of different pinnate phenotypes with haplotypes
Figure BDA0003072045810000082
Note: the backs of commercial roosters could not be analyzed by this method because of only two phenotypes, "-" means no analytical data.
Assisted selection and application of molecular markers:
as shown in Table 5, the H1 haplotype was selected to increase the ratio of black feather in chest and black X black feathered phenotype in back of chest of cock of the F2 generation; the H3 haplotype is selected to improve the chest tingling phenotype, the chest tingling and the feather phenotype of the F2 generation hen, and the chest red feather phenotype, the chest tingling and the feather phenotype proportion of the F2 generation cock; the H2 haplotype can improve the ratio of the yellow feather type in the chest, the yellow feather type in the back and the feather type of jute in the F2 generation hen.
The MC1R gene haplotype can be detected to effectively improve the feather consistency of chicken flocks.
The embodiments in the present description are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Sequence listing
<110> animal science institute of academy of agricultural sciences of Guangdong province
GUANGDONG WIZ AGRICULTURAL SCIENCE & TECHNOLOGY Co.,Ltd.
<120> molecular marker of chicken feather color trait related gene MC1R and application thereof
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cttccccatc cttgtgcctg gggtgcagag gtgcccacat cccctctgcc tcgtgaccgc 60
gtgctgcggg agcactggtg gggctggttg ggcgcacggg ggctttgtag gtgctgcagt 120
tgtgcctcgg ggccacggcc tccagccagg gcggtccctg ggggctgagg ccggggccat 180
gtcgatgctg gcccccctgc gcctcgtgcg cgagccctgg aacgccagtg agggcaacca 240
gagcaatgcc acggccgggg ccggaggtgc ctggtgccag gggctggaca tccccaatga 300
gctcttcctg acgctggggc tggtgagcct ggtggagaac ctgctggtgg tggccgccat 360
cctcaagaac aggaatctgc actcgcccac gtactacttc atctgctgcc tggccgtctc 420
cgacatgctg gtgagcgtca gcaacctggc caagacgctc ttcatgctgc tgatggagca 480
cggcgtgctg gtgatccgcg ccagcatcgt ccgccacatg gacaatgtca tcgacatgct 540
catctgcagc tccgtcgtgt cctccctctc cttcctgggg gtcatcgccg tggaccgcta 600
catcaccatc ttctatgcgc tgcgctacca cagcatcatg acgctgcagc gcgccgtggt 660
caccatggcc agcgtctggc tggccagcac cgtctccagc accgtcttaa tcacctacta 720
ccgcaacaac gccatcctgc tctgcctcat tggcttcttc ctcttcatgc tggtcctcat 780
gctggtgctc tacattcaca tgttcgcgct ggcgtgccac cacgtgcgca gcatctccag 840
ccagcagaag cagcccacca tctaccgcac cagcagcctg aagggagccg tcacgctcac 900
catcctgctg ggagtcttct tcatctgctg ggggcccttc ttcttccacc tcatcctcat 960
cgtcacctgc cccaccaacc ccttctgcac ctgcttcttc agctatttca acctcttcct 1020
catcctcatc atctgcaatt cagtggtcga tcccctgatc tatgccttcc ggagccagga 1080
gctccggcgg acgctgcggg aggtggtgct gtgctcctgg taggaggcgg cacagacagg 1140
aggatggatg gatggatgga tggacggatg gacggatgga tggatggaca aacagatggg 1200
tggatggaca gatgggtgga tggacaaaca gacgcaccgc ggggtgtccc ctgggtgccc 1260
cagtgcagct ggggttgggc tgcctggcct cgcgctccca aataaagg 1308
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cttccccatc cttgtgcctg 20
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<213> Artificial Sequence (Artificial Sequence)
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cctttatttg ggagcgcgag 20

Claims (7)

1. Compared with the MC1R gene coding region DNA sequence SEQ ID NO.1, the molecular marker of the chicken feather color trait related gene MC1R is characterized by comprising the following components: SNP1 at position 69, SNP2 at position 212, SNP3 at position 274, SNP4 at position 644 and SNP5 at position 919;
when the SNP1 is C, SNP2, T, SNP3, G, SNP4 and A, SNP5 which are C, the SNP1 is haplotype H1, and the proportion of the black feather type of the chest and the black X black feather type of the back of the chest of the F2 generation cock is improved;
when the SNP1 is C, SNP2, T, SNP3, G, SNP4 and A, SNP5, the SNP is haplotype H2, and the proportion of a yellow feather type at the chest part, yellow at the back part and a yellow feather type at the jute feather type of the F2 generation hen is improved;
when the SNP1 is T, SNP2, C, SNP3, A, SNP4 and C, SNP5, the SNP is haplotype H3, the proportions of the F2 generation hen chest pockmarked feather type, the back pockmarked feather type and the F2 generation cock chest red feather type, back pockmarked feather type are improved.
2. The molecular marker of the chicken feather trait related gene MC1R as claimed in claim 1, wherein the DNA sequence of coding region of MC1R gene SEQ ID NO: 1, primers used:
a forward primer MC1R-2F: 5'-CTTCCCCATCCTTGTGCCTG-3' as shown in SEQ ID NO. 2;
reverse primer MC1R-2R: 5'-CCTTTATTTGGGAGCGCGAG-3' as shown in SEQ ID NO. 3.
3. The use of the molecular marker of claim 1 in the preparation of a kit for detecting the feathering trait of a chicken.
4. The use of the molecular marker of claim 1 in chicken breeding.
5. A method for detecting molecular markers of chicken feather color-like character related gene MC1R is characterized by comprising the following steps:
(1) performing PCR amplification by using chicken blood DNA as a template;
(2) and carrying out SNP screening on the amplified product to construct a haplotype.
6. The method of claim 5, wherein the PCR amplification system of step (1): in a PCR reaction system with a total volume of 25. mu.L, 1. mu.L of a chicken blood DNA template, 12.5. mu.L of a 2 XPCR reaction mixture, 1. mu.L of each of 10mM forward and reverse primers, 0.25. mu.L of Taq DNA polymerase and 9.25. mu.L double distilled water were added.
7. The method of claim 5, wherein the reaction conditions of the PCR of step (1) are: pre-denaturation at 94 ℃ for 4 min; denaturation at 94 ℃ for 30s, annealing at 62 ℃ for 30s, and extension at 72 ℃ for 1min for 20s, for 34 cycles; finally, the extension is carried out for 10min at 72 ℃ and the product is stored at 4 ℃.
CN202110541670.4A 2021-05-18 2021-05-18 Molecular marker of chicken feather color trait related gene MC1R and application thereof Pending CN113308549A (en)

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Application publication date: 20210827