KR101960427B1 - SNP marker for prediction of dog's tail shape and prediction method using the same - Google Patents

Abstract

The present invention relates to a SNP marker for predicting a shape of dog tails and a prediction method using the same. More specifically, the present invention relates to a composition for predicting the shape of dog tails, a microarray and a kit using the composition, and a method of predicting the shape of dog tails using the same, wherein the shape of curled, hook-like, and rod-like tails can be predicted rapidly and accurately. By utilizing the SNP marker of the present invention, the shape of dog tails can be predicted rapidly and accurately.

Description

SNP marker for predicting tail shape of a dog and a prediction method using the same

The present invention relates to SNP markers for dog tail prediction and a prediction method using the same. More particularly, the present invention relates to SNP markers for dog tail prediction that can quickly and accurately predict the tail shape of a tortoise, a fishing, and a large-sized dog, and a prediction method using the same.

The dog 's tail shape is one of the phenotypic characteristics of the dog' s appearance, and it is necessary to predict early through genetic test because the preference varies according to the tail shape among the dogs.

The tail shape of the dog can be largely classified into a curled type, a fishing type, and a long type. The curled shape means that the tail is substantially completely rounded like a circle. The fishing type means a shape in which the tail is bent to half. In addition, the long and long form means a straight shape without bending the tail.

As a precedent literature on genetic markers for the traits of dogs, "Final Report on the Development of Small Slaughtering by Molecular Sarcoma" Research Institution: Kyungpook National University, organized by the Ministry of Agriculture and Forestry in 2004).

Using the gene tags of the dog's tail, it is possible to meet the preferences of the dogs for the tail shape and to limit the indiscriminate breeding for producing the long or short tail shape, thereby contributing to the protection and welfare policy of the dog. It is anticipated that it will be possible to establish colonial dogs culture and to acquire the scientific value and basic medical information about the body shape of animal by excavating genes related to dog 's tail.

Under these technical backgrounds, the inventors have made intensive efforts to develop gene markers for early prediction of dog tail by extracting genes related to dog tail.

Accordingly, it is an object of the present invention to provide a SNP marker composition for dog tail prediction.

The present invention also provides a method for predicting the dog tail shape using the SNP marker.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to an aspect of the present invention, there is provided a polynucleotide comprising a polynucleotide represented by SEQ ID NO: 1, wherein the 61st base is A or G, and the base sequence of SEQ ID NO: 1 is 5 to 121 consecutive bases There is provided a SNP marker composition for dog tail prediction comprising an agent capable of detecting or amplifying a polynucleotide or its complementary polynucleotide to be constituted.

According to another aspect of the present invention there is provided a kit for dog tail prediction comprising the composition described above.

According to another aspect of the present invention there is provided a microarray for dog tail prediction comprising the composition described above.

According to still another aspect of the present invention, there is provided a method for producing a nucleic acid comprising the steps of: a) extracting nucleic acid from an open sample; And b) identifying the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 1 in the extracted nucleic acid.

According to the embodiment of the present invention, the dog tail can be predicted early, and the effect of being able to satisfy the preference of the dog lover with respect to the dog tail can be achieved.

In addition, it is possible to limit the indiscriminate breeding of dog tail to produce a malt, a fishing, or a large-sized individual, thereby contributing to the protection and welfare policy of the dog, and the colonization of the dog.

Furthermore, by identifying the genes related to the dog 's tail, it is possible to obtain academic value and basic medical information on the animal' s body shape.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Certain terms are hereby defined for convenience in order to facilitate a better understanding of the present invention. Unless otherwise defined herein, scientific and technical terms used in the present invention shall have the meanings commonly understood by one of ordinary skill in the art. Also, unless the context clearly indicates otherwise, the singular form of the term also includes plural forms thereof, and plural forms of the term should be understood as including its singular form.

According to an aspect of the present invention, there is provided a polynucleotide comprising a polynucleotide represented by SEQ ID NO: 1, wherein the 61st base is A or G, and the base sequence of SEQ ID NO: 1 is 5 to 121 consecutive bases There is provided a SNP marker composition for dog tail prediction comprising an agent capable of detecting or amplifying a polynucleotide or its complementary polynucleotide to be constituted.

The SNP markers according to the present invention were found to have high reliability in statistical analysis and tail shape prediction of GWAS results. Therefore, the genotype analysis using the SNP markers can be usefully used as a gene marker capable of early prediction of dog tail.

According to one embodiment of the present invention, the polynucleotide of SEQ ID NO: 2 is A or G at the 61st base, and the base sequence of SEQ ID NO: 2 is 5 to 121 consecutive bases ≪ / RTI > or a complementary polynucleotide thereof; A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 3 and consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 3, or a complementary Polynucleotides; A polynucleotide consisting of 5 to 121 consecutive bases, wherein the 61st base is A or C in the polynucleotide shown in SEQ ID NO: 4 and the 61rd base as the base sequence of SEQ ID NO: 4, or a complementary Polynucleotides; A polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 5 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 5, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 6 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 6, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide shown in SEQ ID NO: 7 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 7, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 8 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 8, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 9 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 9, or a complementary Polynucleotides; The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 10, the polynucleotide consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 10, Polynucleotides; The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 11, the polynucleotide consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 11, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 12 and consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 12, or a complementary Polynucleotides; The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 13, the polynucleotide consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 13, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 14 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 14, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 15 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 15, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 16 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 16, or a complementary Polynucleotides; A polynucleotide having a 61st base of A or C in the polynucleotide shown in SEQ ID NO: 17 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 17 or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 18 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 18 or a complementary Polynucleotides; A polynucleotide having a 61st base of A or G in the polynucleotide shown in SEQ ID NO: 19 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 19, or a complementary Polynucleotides; The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 20, the polynucleotide consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 20, Polynucleotides; And a polynucleotide consisting of 5 to 121 consecutive bases, wherein the 61st base is A or G in the polynucleotide represented by SEQ ID NO: 21 and the 61rd base is the internal base sequence of SEQ ID NO: 21, or a complement thereof Or a polynucleotide capable of detecting or amplifying at least one selected from the group consisting of: < RTI ID = 0.0 > a < / RTI >

According to an embodiment of the present invention, the dog tail may be a curled, a fishing, or a pole.

According to another aspect of the present invention there is provided a kit for dog tail prediction comprising the composition described above.

In the present invention, the kit may be, but is not limited to, an RT-PCR kit or a microarray chip kit including a preparation capable of detecting or amplifying SNP markers.

The RT-PCR kit can comprise a respective pair of primers specific for the marker gene and can be used in combination with other test tubes or other appropriate containers, reagents necessary for PCR amplification, such as buffers, DNA polymerases (e.g., Thermus aquaticus Taq), Thermus thermophilus (Tth), Thermus filiformis, Thermisflavus, Thermococcus literalis or Pyrococcus furiosus (Pfu)), DNA polymerase joins and dNTPs.

The kit may be made from a number of separate packaging or compartments containing the reagent components described above.

According to another aspect of the present invention there is provided a microarray for dog tail prediction comprising the composition described above.

In the present invention, a microarray means a group of polynucleotides immobilized on a substrate at a high density, and the polynucleotide group means a microarray immobilized in a constant region. Such microarrays are well known in the art. The microarrays are described, for example, in U.S. Patent Nos. 5,445,934 and 5,744,305, the contents of which are incorporated herein by reference.

According to still another aspect of the present invention, there is provided a method for producing a nucleic acid comprising the steps of: a) extracting nucleic acid from an open sample; And b) identifying the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 1 in the extracted nucleic acid.

In the present invention, the step of extracting the nucleic acid from the sample of step a) may be performed by a method known in the art. For example, DNA can be directly purified from tissues or cells or amplified by specific amplification of specific regions using amplification methods such as PCR and separation thereof. In the present invention, nucleic acid means not only DNA but also cDNA and RNA molecules synthesized from mRNA. The step of extracting DNA from the target sample can be performed by, for example, PCR amplification, ligase chain reaction (LCR) (Wu and Wallace, Genomics 4, 560 (1989), Landegren et al., Science 241, 1077 USA 86, 1173 (1989)) and self-sustained sequence cloning (Guatelli et al., Proc. Natl. Acad. Sci. USA 87, 1874 (1990)), as well as transcription amplification (Kwoh et al., Proc. Natl. Acad. And nucleic acid-based sequence amplification (NASBA).

According to an embodiment of the present invention, the step of confirming the base type of the SNP of step b) includes alleles specific probe hybridization, allele-specific amplification, Sequence analysis, 5 'nuclease digestion, molecular beacon assay, oligonucleotide ligation assay, size analysis and single strand And a single-stranded conformation polymorphism.

According to an embodiment of the present invention, the amplified gene product may be purified to analyze the base sequence or hybridize with the SNP marker of the present invention.

Methods for amplifying the nucleic acid molecules include PCR, ligase chain reaction, nucleic acid sequence-based amplification, transcription-based amplification system, , Strand displacement amplification or amplification through a Q [beta] replicase, or any other suitable method for amplifying nucleic acid molecules known in the art. Among them, PCR is a method of amplifying a target nucleic acid from a pair of primers that specifically bind to a target nucleic acid using a polymerase. Such PCR methods are well known in the art, and commercially available kits may be used. The PCR can be carried out using a PCR reaction mixture containing various components known in the art necessary for the PCR reaction.

In the present invention, hybridization refers to a process in which two complementary strands of a nucleic acid are combined to form a double stranded molecule (hybrid). In the method of the present invention, the hybridization is carried out under high stringency hybridization conditions.

To detect the degree of hybridization, the target sequence may be labeled with a detectable labeling substance. In one embodiment, the labeling material may be a fluorescent, phosphorescent or radioactive substance, but is not limited thereto. Preferably, the labeling substance is Cy-5 or Cy-3. When the target sequence is amplified, PCR is carried out by labeling the 5'-end of the primer with Cy-5 or Cy-3, and the target sequence may be labeled with a detectable fluorescent labeling substance. In addition, if the radioactive isotope such as P32 or S35 is added to the PCR reaction solution during the PCR, the amplification product may be synthesized and the radioactive substance may be incorporated into the amplification product and the amplification product may be labeled as radioactive.

According to an embodiment of the present invention, the step (b) may include amplifying the extracted nucleic acid.

According to one embodiment of the present invention, further identifying the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 2 to SEQ ID NO: 21.

According to one embodiment of the present invention, in step b), when the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 1 is A, the dog's tail can be predicted as the long tail.

According to one embodiment of the present invention, if one or more of the following cases are selected, the tail of the dog can be predicted as the long tail:

When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 2 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 3 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 4 is C; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 5 is A; The base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 6 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 7 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 8 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 9 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 10 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 11 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 12 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 13 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 14 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 15 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 16 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 17 is A; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 18 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 19 is G; When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 20 is A; And the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 21 is A.

Hereinafter, the present invention will be described in more detail with reference to Examples.

Example  Canine 170K SNP chip analysis for 1. dog

DNA was extracted from the blood of 50 dogs using the Wizard Genomic DNA Purification Kit (Promega, Madison, Wis., USA) and SNP genotyping was performed using CanineHDBeadChip (Illumina, San Diego, CA, USA).

≪ Amplification >

○ Materials and equipment

1. Reagents

2. Organization

Centrifuge, Vortex, Illumina Hybridization oven

○ Experimental Method

1. 20ul of MA1 was dispensed into a MIDI plate (labeled MSA3 plate) with an MSA3 bar code.

2. 4 ul of DNA was placed on the MSA3 plate.

3. Record the DNA ID and the location of the transferred MSA3 plate in the lab tracking form.

4. Add 4 ul of 0.1 N NaOH to each well of MSA3 plate containing MA1 and DNA.

5. Cover the MSA3 plate using a 96-well cap mat and vortex for 1 minute at 1600 rpm.

6. Centrifuge at 280 xg for 1 minute.

7. Reaction was carried out at room temperature for 10 minutes.

8. 34ul MA2 was placed in each well of the MSA3 plate containing the sample.

9. 38ul of MSM was placed in each well of the MSA3 plate containing the sample.

10. Cover the cap mat and centrifuge at 280 x g for 1 min.

11. Reacted in an Illumina Hybridization oven at 37 ° C for 20-24 hours.

(Amplification)

<Day 2 of experiment - Fragment>

○ Materials and equipment

○ Experimental Method

1. Remove the plate from the oven and centrifuge at 50 × g for 1 min.

2. Place 25 ul of FMS into each well containing sample.

3. MSA3 plate was covered with cap mat and vortexed at 1600 rpm for 1 minute.

4. Remove the plate and centrifuge at 50 × g for 1 min.

5. Reaction was carried out in a 37 ° C heat block for 1 hour.

<Day 2 - Precipitation>

○ Materials and equipment

○ Experimental Method

1. Peel the cap mat and place 25 ul of PM1 in each well containing the sample.

2. The cap mat was covered and centrifuged at 1600 rpm for 1 minute.

3. Reaction was carried out at 37 ° C for 5 minutes.

4. Remove the plate and centrifuge at 50 × g for 1 min.

5. Remove the cap mat and place 155 ul of 2-propanol in each well containing the sample.

6. Cover the plate with a new cap mat, turn it 10 times, mix and store at 4 ° C for 30 minutes.

7. Centrifuge at 4 ° C at 3,000 rpm for 20 minutes and immediately remove the MSA3 plate from the centrifuge.

8. Immediately remove the cap mat and quickly flip it over and discard the supernatant.

9. Lightly tapped on absorbent pad (kitchen towel, Kim towel, etc.) 10 times.

10. The inverted plate was placed in a tube rack and air-dried for 1 hour.

<Day 2 - Resuspend >

○ Materials and equipment

○ Experimental Method

1. 23ul RA1 was placed in each well containing DNA pellet and the remaining RA1 was stored for XStain HD Bead Chip (frozen storage).

2. Place a foil seal on the MSA3 plate and seal it by pressing the heat-sealer block for 5 seconds.

3. Reaction was carried out in an Illumina Hybridization oven at 48 ° C for 1 hour

4. Vortexing at 1800 rpm for 1 minute.

5. Centrifuge at 280 xg for 1 minute.

<Day 2 - Hybridization>

○ Materials and equipment

○ Experimental Method

1. MSA3 plates were denatured for 20 minutes in a 95 ° C heat block.

2. After 20 minutes, the MSA3 plate was removed from the heat block and allowed to cool to room temperature for 30 minutes.

3. Nearly 30 minutes to cool the plate, HybChamber was fitted with HybChamber Gaskets.

4. Place 400ul of PB2 in 8 humidifying buffer reservoirs in HybChamber, close HybChamber lid and place at room temperature.

5. After cooling the DNA for 30 min at room temperature, the MSA3 plate was centrifuged at 280 × g for 1 min.

6. Bring the chips from the refrigerator one by one to the chip, unpack the chip package, align the barcode of the HybChamber insert with the barcode of the chip, load the sample on both sides of the chips after 15ul per sample using a multi-channel pipette Respectively.

7. As sample loading of each chip is completed, it is put into HybChamber and the next chip is repeated in the same way.

8. When the chamber is filled, the chamber lid is closed and placed in an Illumina Hybridization oven at 48 ° C, set at a speed of 5, and reacted for 16-24 hours.

<Day 3 - Washing bead chips>

○ Materials and equipment

1. Reagents

2. Organization

- Multi-sample beadChip Alignment fixture

- Te-Flow Flow-Through chambers (black frames, spacers, glass back plate and clamps)

- Wash Dish

- Wash Rack

○ Experimental Method

1. The Hyb chamber was removed from the Hybridization oven.

2. Open the locks of the Hyb chamber and remove them one at a time in the chamber insert.

3. The seal attached to the chip was pulled and removed from the chip.

4. The chip with the thread removed is inserted into the Wash Rack and immersed in the WB1 Wash dish.

5. Once all chips are in the WB1, wash the Rack in a dish for 1 minute, then transfer the Wash Rack to another Wash Dish containing PB1 and repeat this process for 1 minute.

6. After soaking in a PB1 wash dish, the wash rack was removed from the dish for 1 minute, and the wash rack was transferred to another Wash Dish containing PB1 and the process was repeated for 1 minute.

7. After washing, place a back frame on the BeadChips Alignment fixture, place the chips one by one in the direction of the barcode, align the clear space between the white and the alignment to the top and bottom of the alignment fixture I inserted it.

8. Raise the space, place the alignment bar on the top of the chip (no barcode), cover the glass plate with the end of the glass facing the bar, and insert the clip.

(Flow-through chamber assembly completed)

9. After inserting the clip, remove the alignment bar and cut off the space at both ends of the flow-through chamber assembly with scissors.

<Day 3 of the experiment - XStain Beadchips >

○ Materials and equipment

1. Reagents

2. Organization

- Water circulator

- Chamber Rack

- 2 wash dishes, staining rack, tube rack

○ Experimental Method

1. When the temperature of the chamber rack reaches 44 ° C, the flow-through chamber assembly is inserted into the chamber rack.

2. Add 150ul of RA1 to each chip and incubate for 30 seconds. This process was repeated five more times.

3. 450 ul of XC1 was added to each chip and reacted for 10 minutes.

4. Add 450ul of XC2 to each chip and incubate for 10 minutes.

5. Add 200 ul of TEM to each chip and incubate for 10 minutes.

6. 450 ul of 95% formamide / 1 mM EDTA was added to each chip and reacted for 1 minute and then added one more time.

7. Reaction was carried out for 5 minutes.

8. Check the temperature on the label of the LTM tube and change the temperature of the chamber rack according to the temperature.

9. Add 450ul of XC3 to each chip and incubate for 1 minute, then insert again and wait for 8 times to reach temperature.

10. 250 ul of LTM was added to each chip and allowed to react for 10 minutes.

11. Add 450ul of XC3, add one more time after 1 minute, and react for 5 minutes.

12. 250ul ATM was placed in each chip and reacted for 10 minutes.

13. Add 450ul of XC3, add one more time after 1 minute, and react for 5 minutes.

14. 250 ul of LTM was added to each chip and allowed to react for 10 minutes.

15. Add 450ul of XC3, add one more time after one minute, and react for 5 minutes.

16. 250ul ATM was placed in each chip and reacted for 10 minutes.

17. Add 450ul of XC3, add one more time after 1 minute, and react for 5 minutes.

18. 250 ul of LTM was added to each chip and reacted for 10 minutes.

19. Add 450ul of XC3, add one more time after 1 minute, and react for 5 minutes.

20. Upon completion of the procedure, immediately remove the chamber rack from the flow-through chamber, transfer it to the laboratory table at room temperature, and remove it flat.

21. 310 ml of PB1 was placed in the wash container and the dyeing rack was immersed in the container.

22. After removing the clips of the chamber rack and lifting the glass block with the aid of the apparatus, the spaces where the beads of the chips are stuck on both ends were removed.

23. Once all the attachments on the chip have been removed, they are inserted into the staining rack contained in PB1 and immersed in PB1. All chips were processed in the same way.

24. Slowly move the dye rack up and down about 10 times to quench the chips and soak them for 5 minutes.

25. Place 310 ml of XC4 in another container for cleaning, quench 10 times in the same manner as 24, and soak for 5 minutes.

26. After 5 minutes, the dyeing rack was removed from the cleaning vessel and placed on a tube rack.

27. Using the forceps, carefully remove the chip from the rack and place it on the tube rack.

28. The tube rack on which the chips were placed was carefully placed in a vacuum dryer and allowed to dry for 55-55 minutes under a vacuum of 508 mm Hg (0.68 bar).

29. Once it was confirmed that the chip was dried, the edge of the chip was closed well using Kim wipes soaked in ethanol. At this time, the bead portion was careful not to touch.

30. The bead chip was imaged using a scanner within 72 hours after completion of the experiment.

result

Twenty-one SNPs with predictable traits were selected from 50 high-density SNP 170K chip experiments and full-length genome association analysis (GWAS) (Table 1), and the SNP nucleotide sequence information is shown in Table 2 below . Table 1 shows the results of statistical analysis of 21 single base mutations (SNPs) capable of early prediction of dog tails. Table 2 shows the nucleotide sequence information of SNPs that can predict early canine tail.

Table 3 shows the mean and standard error values of the genotypes associated with dog tail (① curling type, ② fishing type, and ③ long size) by box plot.

In Table 3, the dog type is a number 1, the fishing type (middle type) is a number 2, and the long type (straight type) is a number 3 in the dog tail. The first SNP with the highest statistical significance is AA, AG, and GG, and the number of dogs corresponding to this number is 2, 15, and 33, respectively, as shown in the figure inside. In this case, the mean value of genotype-1 is 2.5, which means that the average of two tails with AA genotype has a median value of 2 and 3. Similarly, genotype-2 AG genotypes have 15 of these genotypes and their average number of tails is 2.27. Genotype-3 has 33 GG genotypes and their average value is 1.58. This means that 33 of the tails have an intermediate shape between the tails and the tines.

The present invention selects 21 SNPs that are significantly associated with dog tail through high density SNP 170K chip experiment and GWAS analysis. These SNPs can be used as a genetic marker and can be used as a genetic marker for early prediction of dog tail shape.

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the present invention is not limited thereby. something to do. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

<110> RURAL DEVELOPMENT ADMINISTRATION <120> SNP marker for prediction of dog's tail shape and prediction          method using the same <130> NPF31610 <160> 21 <170> PatentIn version 3.2 <210> 1 <211> 121 <212> DNA <213> Canis familiaris <400> 1 tggatggata gagcagaagc aagccctgct tgctagagac tagacacatc actggaacct 60 accatgtcta agtgcaggtg gcccaaggga aggagacagg gcataytgcc ttgcttgtgt 120 a 121 <210> 2 <211> 121 <212> DNA <213> Canis familiaris <400> 2 taactcaccg gctcgctgtg tgtgtagtag gcaacactcc taaaaggttt ctcacggggg 60 aacggggtca cctatgcctg caaattgctc aaaattcact tttgaatgag ctcctattag 120 a 121 <210> 3 <211> 121 <212> DNA <213> Canis familiaris <400> 3 gggaagggag ttgaggttgc cctggcactc cttcagccac tgcatcctgc tcaggactgc 60 atgggacttg ggtgtgaaca ccctgcatcc agctcctgtg attgagagcg atccttccca 120 a 121 <210> 4 <211> 121 <212> DNA <213> Canis familiaris <400> 4 tcggggtgcc ctgatgccac tgagcccctt tcatacccac tcctcattag aacatattct 60 actttttctc cattgtgaga gagataatat agcagtgttc tgttaatagt gggatagtca 120 c 121 <210> 5 <211> 121 <212> DNA <213> Canis familiaris <400> 5 ctttaagaac tgtgtgaatt catgacaaga tatttacatt tggactattt agagcagagt 60 ataagagaga ctacacaaac caaagttcag agatggagcc attcctggag acctgtgtag 120 t 121 <210> 6 <211> 121 <212> DNA <213> Canis familiaris <400> 6 atgagtgtgc attgagtact gcttcttcca tgtggctagc tctgtgataa gcaccatgaa 60 actttaaaaa atggtgagaa ttactttaag aactgtgtga attcatgaca agatatttac 120 a 121 <210> 7 <211> 121 <212> DNA <213> Canis familiaris <400> 7 aacacatgac aagtgtgaga gcaaaatggt tcccaaagtg cacagctttg tataaaaacc 60 agcagcataa gaagctgcca ttacatacat atttcctgaa aaacatatct agcttagtga 120 g 121 <210> 8 <211> 121 <212> DNA <213> Canis familiaris <400> 8 cctcaagagg acatgtctgt gctactggac ttgaaaagaa ggaagggagt atccaagtac 60 aaagggaagg ggaagagaaa ctccaagttt ccctgagaag gtggcattcc aagaagagat 120 g 121 <210> 9 <211> 121 <212> DNA <213> Canis familiaris <400> 9 acactacacc cgatataatg atatgtcatt tcaatgtagc tagcccatga acttctacag 60 atttaatggg attttgcgta tttagaattt catcataaag gggccctttg gaatcattat 120 a 121 <210> 10 <211> 121 <212> DNA <213> Canis familiaris <400> 10 gtccccgcac cagcctcagg gcccgtcacc aggcgcccgt gccccctcgc tccccggcac 60 agtctctctt cttcacgaac cagccagccc gacggtctga gcttccgatg tcctcctggg 120 c 121 <210> 11 <211> 121 <212> DNA <213> Canis familiaris <400> 11 tctaactaga gtattccttc agagcctcag agaacacaga cattttccca agattacaga 60 asaagacaca ccaaggctgc aacttatgtg tcttgattct gggtccagtg ctccatgyac 120 a 121 <210> 12 <211> 121 <212> DNA <213> Canis familiaris <400> 12 atgcaagtat aactgaatac tgtacaatta tttaatttta aactatttta caggatagct 60 accttcamta tttagtgtac tgcctgggat gcagtaatgc taaatatcat gttagacctc 120 c 121 <210> 13 <211> 121 <212> DNA <213> Canis familiaris <400> 13 tagttctgaa agaagtcaaa caaattttga acttccatct tgggttttgt aagtaatgtg 60 atccatatag acttgtgatg aaattggagt ggtaaactct tccaaaacac catatyggta 120 a 121 <210> 14 <211> 121 <212> DNA <213> Canis familiaris <400> 14 cccgaatgat ttagcagcca tgggaaaaga aaaggaatgt tacaataaat tatgtaataa 60 agaaatgata tacaaagcaa catattgtgc gaagggtggc ctttagcttc cacacttctt 120 g 121 <210> 15 <211> 121 <212> DNA <213> Canis familiaris <400> 15 gtggtgtttg ccggggatgt ggagcagcgc cgagttctgc cctgcaaacg catctcgaat 60 aatgaaagct ggtcttccac tttgcagcaa gcctttaagc cacttcgaag actactttac 120 t 121 <210> 16 <211> 121 <212> DNA <213> Canis familiaris <400> 16 tacctcccat ctgagacaat taaaggtggg tgtggatttt gtatggtctc ttgttttcct 60 atagattgga tataaagatc cccaagttgg caaggtacaa gatagaagca ttctaggtcc 120 a 121 <210> 17 <211> 121 <212> DNA <213> Canis familiaris <400> 17 aaaaagagca agatggtgat attgctcaga acactaaagc acccctatgt cagtgtgctc 60 agcaaattag ccagtgttac ttgtattgcc ataaaaaaaa cacacacaca ttttaaggaa 120 g 121 <210> 18 <211> 121 <212> DNA <213> Canis familiaris <400> 18 agaagtttcc taggttgagt ttggaaggta agtacatttg ccagagagac aggaagagag 60 aaagaaaagt ggaagaggaa aacagaagcc ggcctccttt agggaccata gcatgcaagt 120 g 121 <210> 19 <211> 121 <212> DNA <213> Canis familiaris <400> 19 aatgatactc tgcagcccct gcattaataa aagaatggaa aaaaaaaaaa aaaagaatgg 60 actctgattt cagacagtct aatgagtttg aaattacctc aaaatacagg atactaccta 120 g 121 <210> 20 <211> 121 <212> DNA <213> Canis familiaris <400> 20 tacaggcccc aggaaggagc caccagatgc ccaggactgg gcccaggaat gatggaggct 60 atacagctgg ctgcctgcac tggctgccgc ccctgtcatc cagtgtcaca gagcagcacc 120 t 121 <210> 21 <211> 121 <212> DNA <213> Canis familiaris <400> 21 gggaagggag ttgaggttgc cctggcactc cttcagccac tgcatcctgc tcaggactgc 60 atgggacttg ggtgtgaaca ccctgcatcc agctcctgtg attgagagcg atccttccca 120 a 121

Claims (10)

A polynucleotide consisting of 5 to 121 consecutive bases, wherein the 61st base is A or G in the polynucleotide represented by SEQ ID NO: 1, and the 61rd base is an internal nucleotide sequence of SEQ ID NO: 1, or a complementary A composition for dog tail prediction, comprising an agent capable of detecting or amplifying a polynucleotide,
Wherein the dog tail shape is a curled, a fishing, or an elongated shape. The method according to claim 1,
A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 2 and consisting of 5 to 121 consecutive bases containing the 61st base as the nucleotide sequence of SEQ ID NO: 2, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 3 and consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 3, or a complementary Polynucleotides;
A polynucleotide consisting of 5 to 121 consecutive bases, wherein the 61st base is A or C in the polynucleotide shown in SEQ ID NO: 4 and the 61rd base as the base sequence of SEQ ID NO: 4, or a complementary Polynucleotides;
A polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 5 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 5, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 6 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 6, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide shown in SEQ ID NO: 7 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 7, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 8 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 8, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 9 and consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 9, or a complementary Polynucleotides;
The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 10, the polynucleotide consisting of 5 to 121 consecutive bases including the 61st base as the base sequence of SEQ ID NO: 10, Polynucleotides;
The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 11, the polynucleotide consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 11, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 12 and consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 12, or a complementary Polynucleotides;
The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 13, the polynucleotide consisting of 5 to 121 consecutive bases containing the 61st base as the base sequence of SEQ ID NO: 13, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 14 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 14, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide of SEQ ID NO: 15 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 15, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 16 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 16, or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or C in the polynucleotide shown in SEQ ID NO: 17 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 17 or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 18 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 18 or a complementary Polynucleotides;
A polynucleotide having a 61st base of A or G in the polynucleotide shown in SEQ ID NO: 19 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 19, or a complementary Polynucleotides;
The polynucleotide having the 61st base of A or G in the polynucleotide shown in SEQ ID NO: 20, the polynucleotide consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 20, Polynucleotides; And
A polynucleotide having a 61st base of A or G in the polynucleotide represented by SEQ ID NO: 21 and consisting of 5 to 121 consecutive bases including the 61st base as the nucleotide sequence of SEQ ID NO: 21, or a complementary A polynucleotide; and an agent capable of detecting or amplifying at least one selected from the group consisting of polynucleotides. delete A kit for dog tail prediction comprising the composition according to any one of claims 1 or 2. 4. A microarray for predicting the tail of a dog comprising the composition according to any one of claims 1 or 2. a) extracting a nucleic acid from an open sample; And
b) identifying the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 1 in the extracted nucleic acid,
Wherein in step b), if the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 1 is A, the dog tail is predicted as the long tail. The method according to claim 6,
Wherein the base type of the SNP corresponding to the 61st base of the at least one polynucleotide of SEQ ID NO: 2 to SEQ ID NO: 21 is further confirmed. The method according to claim 6,
And amplifying the extracted nucleic acid in step b). delete 8. The method of claim 7,
Predicting a dog's tail by predicting the dog's tail as a long-tail if it corresponds to one or more of the following cases:
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 2 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 3 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 4 is C;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 5 is A;
The base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 6 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 7 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 8 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 9 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 10 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 11 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 12 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 13 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 14 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 15 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 16 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 17 is A;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 18 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 19 is G;
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 20 is A; And
When the base type of the SNP corresponding to the 61st base of the polynucleotide of SEQ ID NO: 21 is A.