KR20220066755A - Single nucleotide polymorphisms associated with heat resistance of African indicine breeds and their application - Google Patents

Abstract

The present invention relates to a heat resistance African humped cattle (AFH) variety specific single nucleotide polymorphism and a use thereof. A heat resistance AFH variety sorting kit according to the present invention can detect individuals having enhanced heat resistance in AFH varieties with high accuracy, thereby having an advantage of being variously used for reproductive gene research based on African cattle genome research and gene analysis.

Description

Single nucleotide polymorphisms associated with heat resistance of African indicine breeds and their application}

The present invention relates to heat resistant cattle African Humped Cattle (AFH) breed specific mononucleotide polymorphisms and uses thereof.

Cattle are an important economically and socially important resource in Africa. About 150 endemic cattle breeds inhabit sub-Saharan Africa. For a long time, native African cattle living in isolated areas have been selected for environmental factors. This acted as a strong selective pressure on African cattle and led to selection of individuals more adapted to the harsh environment. In particular, some cultivars have acquired resistance to lethal local diseases that reduce survival and productivity. In addition to environmental factors, selection from humans also influenced phenotypic traits that improved production of dairy and meat quality.

Large-scale genetic mutations that occurred rapidly in a short period of time made it possible to identify the location of genes or loci that control expression traits. Whole genome analysis ultimately allows us to understand the role of unique genetic signatures in adaptation to the environment. Several recent genomic analyzes have been performed to study the diversity and genetic background of various African cattle breeds. For example, SNP analysis of small African zebu to improve livestock yields enabled discovery of candidate loci associated with the trait. The discovery of these genetic loci showed genetic diversity related to the phenotypic traits of a specific breed, and made it possible to interpret the genome functionally.

On the other hand, most cattle in Africa today have B. taurus species and B. indicus species as their ancestors, and it is known that they are B. taurus x B. indicus individuals of various phenotypes. African humped cattle have only taurine mtDNA haplotype, and Y chromosome microsatellite has both indicine and taurine Y chromosomes in Africa. Furthermore, autosomal full-length genome analysis shows that African cattle contain taurine genes with different genetic contributions across populations, but show little variation within populations, suggesting that natural selection in African cattle is B. taurus x B. indicus played an important role in shaping the mixing ratio, suggesting that genetic mixing increased diversity and provided new genetic resources for humans and natural selection.

Against this background, studies are being attempted to discover genetic markers that can maintain high survival and productivity in African cattle through taurine or indicine-derived gene diversity analysis.

The present inventors tried to identify a heat-resistant African Humped Cattle (AFH) breed-specific genetic signature that exhibits a high survival rate in harsh environmental conditions. As a result, the present invention was completed by identifying SNPs specific to heat-resistant cattle AFH through comparative genome-wide analysis of cattle of various breeds.

Accordingly, an object of the present invention is to provide a kit for screening heat-resistant cattle African Humped Cattle (AFH) varieties.

Another object of the present invention is to provide a method for detecting a heat resistant bovine AFH breed-specific marker.

The present invention relates to a heat-resistant cattle African Humped Cattle (AFH) breed-specific mononucleotide polymorphism and its use, and the kit for screening heat-resistant cattle AFH breeds according to the present invention is an individual with enhanced heat resistance in AFH breeds with high accuracy. can be detected.

Hereinafter, the present invention will be described in more detail.

One aspect of the present invention is a single nucleotide polymorphism (SNP) site of the DNAJC18 gene downstream (Downstream), single nucleotide polymorphism sites at positions 922, 6511 and 7378 of SEQ ID NO: 1;

single nucleotide polymorphism (SNP) sites of the DNAJC18 gene, single nucleotide polymorphism sites at positions 12439, 14758, 16701, 25415, 28427, 29222, 30021 and 30051 of SEQ ID NO: 2; and

As a single nucleotide polymorphism (SNP) site of the DNAJC18 gene upstream, 1193, 1301, 2072, 2073, 2588, 2971, 4246, 4319, 4507, 4521, 4626, 4794, 4902, 6367, single nucleotide polymorphism sites at positions 7164, 7603, 7971, 8149 and 8301;

It is a kit for screening heat-resistant AHF (African Humped Cattle) breeds comprising a primer or probe that specifically binds to 10-100 consecutive nucleotide sequences including at least one single nucleotide polymorphism site.

In one embodiment of the present invention, the AFH variety may be a kit having a single nucleotide polymorphism site having a T at nucleotide position 922 of SEQ ID NO: 1.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 6511 of SEQ ID NO: 1 .

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 7378 nucleotide position of SEQ ID NO: 1.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 12439th nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 14758th nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having T at the 16701 nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having T at the 25415th nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 28427th nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 29222 nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 30021 of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 30051 nucleotide position of SEQ ID NO:2.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 1193 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at the 1301 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 2072 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at the 2073 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 2588th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 2971 of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having C at the 4246th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 4319 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 4507 of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 4521 of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 4626th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit having a single nucleotide polymorphism site having a G at the 4794 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 4902 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having A at nucleotide position 6367 of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having G at the 7164th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a G at the 7603 nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 7971th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 8149th nucleotide position of SEQ ID NO:3.

In one embodiment of the present invention, the AFH variety may be a kit, which has a single nucleotide polymorphism site having a T at the 8301 nucleotide position of SEQ ID NO:3.

As used herein, the term “nucleotide” refers to deoxyribonucleotides or ribonucleotides that exist in single-stranded or double-stranded form, and unless otherwise specified, includes analogs of natural nucleotides.

As used herein, the term “single nucleotide polymorphism (SNP)” occurs when a single nucleotide (A, T, C, or G) in the genome differs between members of a species or between pairs of chromosomes of an individual. refers to the diversity of DNA sequences. For example, when three DNA fragments from different individuals (eg, AAGT[A/A]AG, AAGT[A/G]AG, AAGT[G/G]AG) contain differences in a single base, Called two alleles (A or G), in general almost all SNPs have two alleles. Within a population, SNPs can be assigned a minor allele frequency (MAF; the lowest allele frequency at a locus found in a particular population). Variations exist within human populations, and single SNP alleles common in geological or ethnic groups are very rare.

A single nucleotide polymorphism may be included in the coding sequence of a gene, in a non-coding region of a gene, or in a region between genes. SNPs in the coding sequence of genes do not necessarily cause changes in the amino acid sequence of the target protein due to the degeneracy of the genetic code. SNPs that form the same polypeptide sequence are said to be synonymous (also called silent mutations), and SNPs that form different polypeptide sequences are said to be non-synonymous. Non-synonymous SNPs can be missense or nonsense, where a missense change results in another amino acid while a nonsense change forms an immature stop codon. SNPs that are not present in protein-coding sites can cause gene silencing, transcription factor binding, or non-coding RNA sequences.

As used herein, the term “primer” refers to an oligonucleotide, and conditions in which the synthesis of a primer extension product complementary to a nucleic acid chain (template) is induced, that is, the presence of nucleotides and a polymerization agent such as DNA polymerase, and a suitable temperature It can act as a starting point for synthesis under the conditions of and pH. For example, a primer is a deoxyribonucleotide and is single-stranded. The primer used in the present invention may include naturally occurring dNMP (ie, dAMP, dGMP, dCMP, and dTMP), modified nucleotides, or non-natural nucleotides. In addition, the primer may also include ribonucleotides.

The primer of the present invention may be an extension primer that is annealed to a target nucleic acid to form a sequence complementary to the target nucleic acid by a template-dependent nucleic acid polymerase, which extends to the position where the immobilized probe is annealed so that the probe is It occupies the annealed area.

The extension primer used in the present invention includes a hybridization nucleotide sequence complementary to the first position of the target nucleic acid. The term “complementary” means that a primer or probe is sufficiently complementary to selectively hybridize to a target nucleic acid sequence under certain annealing or hybridization conditions, and is substantially complementary and perfectly complementary. It has an all-inclusive meaning, for example, completely complementary. In the present specification, the term “substantially complementary sequence” used in relation to a primer sequence means not only a completely identical sequence, but also a sequence to be compared partially within the range that can function as a primer by annealing to a specific sequence. It is meant that sequences that do not match are also included.

The primer should be long enough to prime the synthesis of extension products in the presence of a polymerizer. A suitable length of a primer depends on a number of factors, such as temperature, application, and source of the primer, but is typically 15-30 nucleotides. Short primer molecules generally require lower temperatures to form sufficiently stable hybrid complexes with the template. The term “annealing” or “priming” refers to the apposition of an oligodeoxynucleotide or nucleic acid to a template nucleic acid, wherein the apposition is a polymerase that polymerizes the nucleotides to form a nucleic acid molecule complementary to the template nucleic acid or a portion thereof. make it

The sequence of the primer does not need to have a completely complementary sequence to a partial sequence of the template, but it is sufficient if it hybridizes with the template and has sufficient complementarity within a range capable of performing an intrinsic action of the primer. Therefore, the primer in the present invention does not need to have a perfectly complementary sequence to the above-described nucleotide sequence as a template, and it is sufficient if it hybridizes to this gene sequence and has sufficient complementarity within the range to act as a primer. The design of such a primer can be easily performed by those skilled in the art with reference to the above-described nucleotide sequence, for example, using a primer design program (eg, PRIMER 3 program).

As used herein, the term “nucleic acid molecule” has a meaning comprehensively including DNA (gDNA and cDNA) and RNA molecules, and the nucleotide, which is a basic structural unit in a nucleic acid molecule, is a natural nucleotide as well as a sugar or base site modified Also includes analogs that have been

When the starting material in the kit of the present invention is gDNA, isolation of gDNA may be performed according to a conventional method known in the art.

In the kit of the present invention, identifying the specific sequence can be performed by applying various methods known in the art. For example, techniques applicable to the present invention include fluorescence in situ hybridization (FISH), direct DNA sequencing, PFGE analysis, Southern blot analysis, single-stranded conformation analysis, RNase protection assay, dot blot analysis, denaturation. gradient gel electrophoresis, methods using proteins that recognize nucleotide mismatches, and allele-specific PCR.

Sequence changes result in differences in base binding within single-stranded molecules, resulting in the appearance of bands with different mobilities, which SSCA detects. DGGE analysis uses a denaturing gradient gel to detect sequences that exhibit different mobility than wild-type sequences.

Other techniques generally use probes or primers complementary to sequences comprising the SNPs of the invention.

For example, in an RNase protection assay, a riboprobe complementary to a sequence comprising SNPs of the invention is used. The riboprobe and DNA or mRNA isolated from humans are hybridized, and then digested with an RNase A enzyme capable of detecting a mismatch. If there is a mismatch and RNase A recognizes it, a smaller band is observed.

In the analysis using the hybridization signal, a probe complementary to the sequence comprising the SNP of the present invention is used. In this technique, the risk of disease is directly determined by detecting the hybridization signal of the probe and the target sequence.

As used herein, the term “probe” refers to a natural or modified monomer including deoxyribonucleotides and ribonucleotides capable of hybridizing to a specific nucleotide sequence or a linear oligomer having a bond. For example, the probe is single-stranded for maximum efficiency in hybridization. The probe is preferably a deoxyribonucleotide.

As the probe used in the present invention, a sequence that is completely complementary to the sequence including the SNP may be used, but a sequence that is substantially complementary within a range that does not interfere with specific hybridization may be used. may be For example, the probe used in the present invention comprises a sequence capable of hybridizing to a sequence comprising 10-30 contiguous nucleotide residues comprising a SNP of the present invention. More preferably, the 3'-end or 5'-end of the probe has a base complementary to the SNP base. In general, since the stability of the duplex formed by hybridization tends to be determined by the match of the sequences at the ends, probes having a base complementary to the SNP base at the 3'-end or the 5'-end end If the parts do not hybridize, these duplexes can disintegrate under stringent conditions.

Stringent conditions used for hybridization may be determined by controlling temperature, ionic strength (buffer concentration), and the presence of compounds such as organic solvents. These stringent conditions may be determined differently depending on the sequence to be hybridized.

Another aspect of the present invention is in a biological sample of cattle to provide information necessary for the selection of heat-resistant cattle AHF (African Humped cattle) breeds,

A single nucleotide polymorphism (SNP) site of the DNAJC18 gene downstream, single nucleotide polymorphism sites at positions 922, 6511 and 7378 of SEQ ID NO: 1;

single nucleotide polymorphism (SNP) sites of the DNAJC18 gene, single nucleotide polymorphism sites at positions 12439, 14758, 16701, 25415, 28427, 29222, 30021 and 30051 of SEQ ID NO: 2; and

As a single nucleotide polymorphism (SNP) site of the DNAJC18 gene upstream, 1193, 1301, 2072, 2073, 2588, 2971, 4246, 4319, 4507, 4521, 4626, 4794, 4902, 6367, single nucleotide polymorphism sites at positions 7164, 7603, 7971, 8149 and 8301;

It is a method of detecting a heat-resistant cattle AHF (African Humped cattle) breed-specific marker comprising the step of detecting one or more single nucleotide polymorphism sites among them.

As used herein, the term "biological sample" refers to any material of an organism, including nucleic acids, and the biological samples that can be used in the nucleic acid amplification reaction in the present invention include viruses, bacteria, tissues, cells, hair, oral tissues, oral cells, and blood. , lymph, bone marrow fluid, saliva, milk, urine, feces, ocular fluid, semen, brain extract, spinal fluid, joint fluid, mammary fluid, ascites, amniotic fluid, or tissue fluid. As a biological sample, blood includes whole blood, plasma, or serum.

In one embodiment of the present invention, the biological sample is whole blood, plasma or serum.

The step of detecting the single nucleotide polymorphism (SNP) site of the present invention may be performed in a microarray method or a gene amplification method.

In the present invention, when the detection step is performed by a gene amplification method, it may be performed according to polymerase chain reaction (PCR) using primers.

In one embodiment of the present invention, the PCR process is reverse transcription-polymerase chain reaction (RT-PCR), reverse transcription-quantitative PCR (RT-qPCR), real-time nested reverse transcription- Polymerase chain reaction (realtime nested RT-PCR; rt-nRT-PCR) or digital reverse transcription polymerase chain reaction (digital RT-PCR; dRT-PCR) may be performed, but is not limited thereto.

When the method of the present invention is carried out using primers, the nucleotide sequence of the marker of the present invention is analyzed and confirmed by conducting a gene amplification reaction. Since the present invention detects the nucleotide sequence of the marker of the present invention, by determining the nucleotide sequence of the marker of the present invention in the sample to be analyzed (eg, genomic DNA), it is possible to confirm whether the target sample is an AFH variety.

In one embodiment of the present invention, the method may be implemented in a microarray manner. When the method of the present invention is a microarray method, a probe may be immobilized on a solid surface of the microarray.

The probe used in the method of the present invention may have a sequence complementary to a sequence of 10-100 consecutive nucleotides on each gene including the SNPs of the present invention listed above.

In the microarray of the present invention, the probe is used as a hybridizable array element and is immobilized on a substrate. Preferred gases include suitable rigid or semi-rigid supports, such as membranes, filters, chips, slides, wafers, fibers, magnetic or non-magnetic beads, gels, tubing, plates, polymers, microparticles and capillaries. The hybridization array elements described above are arranged and immobilized on the substrate. Such immobilization is carried out by a chemical bonding method or a covalent bonding method such as UV. For example, the hybridization array element may be bonded to a glass surface modified to include an epoxy compound or an aldehyde group, and may also be bonded by UV to a polylysine coated surface. In addition, the hybridization array element may be bound to a gas through a linker (eg, ethylene glycol oligomer and diamine).

The sample DNA applied to the microarray of the present invention may be labeled and may be hybridized with an array element on the microarray. Hybridization conditions may vary. Detection and analysis of the degree of hybridization may be variously performed depending on the labeling material.

The label of the probe may provide a signal to detect hybridization, which may be linked to an oligonucleotide. Suitable labels include fluorophores (eg, fluorescein, phycoerythrin, rhodamine, lissamine, and Cy3 and Cy5 (Pharmacia)), chromophores, chemiluminophores, magnetic particles, radioisotopes. Elements (P32 and S35), mass labels, electron-dense particles, enzymes (alkaline phosphatase or horseradish peroxidase), cofactors, substrates for enzymes, heavy metals (eg gold) and antibodies, streptavidin, biotin , digoxigenin and haptens having specific binding partners such as chelating groups, but are not limited thereto. Labeling can be carried out through various methods commonly practiced in the art, such as nick translat (bio method, random priming method, and kynation method). Labeling can be performed by fluorescence, radioactivity, chromometry, gravimetric measurement, X -Provides a detectable signal by ray diffraction or absorption, magnetic, enzymatic activity, mass analysis, binding affinity, hybridization radiofrequency, and nanocrystals.

In the present invention, a nucleic acid sample to be analyzed can be prepared using mRNA obtained from various biosamples. A hybridization reaction-based analysis may be performed by labeling the cDNA to be analyzed instead of the probe.

When a probe is used, the probe can be hybridized with a cDNA molecule. In the present invention, suitable hybridization conditions may be determined in a series of processes by an optimization procedure. These procedures are carried out as a series of procedures by those skilled in the art to establish protocols for use in the laboratory. For example, conditions such as temperature, concentration of components, hybridization and washing times, buffer components and their pH and ionic strength may depend on various factors such as the length of the probe and the amount of GC and the target nucleotide sequence.

After the hybridization reaction, a hybridization signal emitted through the hybridization reaction is detected. The hybridization signal can be carried out in various ways, for example, depending on the type of label bound to the probe. For example, when the probe is labeled with an enzyme, hybridization may be confirmed by reacting a substrate of the enzyme with a hybridization reaction product. Enzyme/substrate combinations that can be used include peroxidase (eg horseradish peroxidase) with chloronaphthol, aminoethylcarbazole, diamino benzidine, D-luciferin, lucigenin (bis-N-methylacridinium). nitrate), resorufin benzyl ether, luminol, Amplex Red reagent (10-acetyl-3,7-dihydroxyphenoxazine), HYR (p-phenylenediamine-HCl and pyrocatecN-l), TMB (tetramethyl benzidine), ABTS (2,2-Azino-di-[3-ethylbenzthiazoline sulfate]), o-phenylenediamine (OPD minoethylnaphthol/pyronine; alkaline phosphatase and bromochloroindolyl phosphate (BCIP), nitro blue tetrazolium (NBT), naphthol-AS-B1-phosphate (naphtN-l-AS-B1-pN-spNate) noethylECF substrate; glucose oxidase and tNBT (nitroblue tetrazolium) and m-PMS (phenzaine methosulfate). When the probe is labeled with gold particles, it can be detected by a silver staining method using silver nitrate.Therefore, when the method for detecting the marker of the present invention is carried out based on hybridization, specifically (i) the present invention Hybridizing a probe having a sequence complementary to the nucleotide sequence of the marker to a nucleic acid sample (ii) detecting whether the hybridization reaction occurs. By analyzing the intensity of the hybridization signal by the hybridization process, AFH It is possible to determine whether a variety exists, that is, when a hybridization signal to the nucleotide sequence of the marker of the present invention is stronger than that of a normal sample, it is determined as a heat-resistant AFH variety.

The present invention relates to a heat-resistant cattle African Humped Cattle (AFH) breed-specific mononucleotide polymorphism and its use, and the kit for screening heat-resistant cattle AFH breeds according to the present invention is heat-resistant in AFH breeds with high accuracy. Because it can detect enhanced individuals, it has the advantage of being able to be used in a variety of ways in African cattle genome research and reproductive gene research based on gene analysis.

1 is a graph showing the results of an integrated haplotype score (iHS) analysis of AFH varieties according to an embodiment of the present invention.
2 is a graph showing the analysis results of the iHS taurine ancestor of the AFH variety according to an embodiment of the present invention.
3A is a graph showing the results of analyzing candidates for the selective locus on BTA11 in which the gene of the indicine ancestor is excessively expressed according to an embodiment of the present invention.
3A is a heat map showing the results of analyzing candidates for the selective locus on BTA11 in which the gene of the indicine ancestor is excessively expressed according to an embodiment of the present invention.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, these examples are only for illustrating the present invention, and the scope of the present invention is not limited by these examples.

Experimental Example 1: Gene sequencing

A total of 116 African cattle genome samples were sequenced according to the TruSeq DNA Sample Prep Kit v.2 Support (FC121-2001). After fragmenting 1 μg of genomic DNA using a Covaris Focused-Ultrasonicator, an 'A' sequence and an adapter sequence were added to the 3' end. Among the fragmented DNA fragments, fragments with a length of 400 to 500 bp were selected, and after PCR amplification, a quality verification process using Bioanalyzer was performed, and then the selected DNA fragments were sequenced using HiSeq2000.

In addition to the above sequencing data, a total of 53 commercial taurine breeds, 48 African cattle, and 10 African taurines, 50 European taurines, and 34 American- Australian zebu and 22 Asian Zebu varieties were used.

Genetic information was processed according to the guidelines provided by the 1000 Bull Genome Project as of October 17, 2019. The quality of the genomic data produced was confirmed using the fastQC (v.0.11.8) program, and sequences and artifacts with low quality were removed using the Trimmomatic (v.0.39) program. The genome fragment information obtained through this was aligned to the reference genome ARS-UCD1.2 using the default parameters of bwa mem (v.0.7.17), and further processed using Samtools (v.1.9). Duplicate sequences generated by PCR were found using the 'MarkDuplicates' option of the Picard v.2.20.2 program, and the quality scores of the sequences were corrected using the 'BaseRecalibrator' option of GATK (v.3.8). Data produced by the 1000 Bull Genome Project except for two African buffalo were used as mutation information in cattle. Validation of results before and after the procedure for sequence quality score correction (base quality score recalibration; BQSR) was done through the 'AnalyzeCovariates' option. The procedure for correcting the sequence quality score was repeated using the mutations found after analysis.

Experimental Example 2: Gene mutation discovery

To discover genetic mutation candidates, GVCF files were created using the 'HaplotypeCaller' and '-ERC GVCF' options of the GATK program, and the GVCF files created for each individual were combined using the 'CombineGVCFs' option of the GATK program. The combined GVCF files were used to discover single nucleotide polymorphisms (SNPs), one of the representative genetic variants, through the 'GenotypeGVCFs' and 'SelectVariants' options. At this time, the method suggested by GATK was referred to minimize the type I error of variation information: (1) SNP grouped in '-clusterSize 3', '-clusterWindowSize 10', (2) Average DP < 1/3X and > 3X, (3) QD < 2, (4) QUAL < 30, (5) SOR >3 (6) FS > 60, (7) MQ < 40, (8) MQRankSum < -12.5, (9) ReadPosRankSum < - SNPs corresponding to 8 were removed. In addition, SNPs having two or more alleles and SNPs missing genotype information in more than 99% of samples were also removed. The SNPs left through this were subjected to imputation and phasing using the BEAGLE 4.0 (r1399) program. Finally, SNPs with less than 0.01% of all genotypes with a low frequency of alleles were additionally filtered, and the remaining SNPs was annotated using the SNPEff (v.4.3) program.

To confirm the reliability of genetic variation information, it was compared with the genotype information of 69 individuals obtained through the BovineSNP50 Genotyping BeadChip (Illumina). GeneCall score excluding SNPs of less than 0.7 were compared, and the reliability of the genetic variation discovery pipeline of this study was confirmed by using the genotype data of 45 individuals used in the previous study.

Experimental Example 3: Excavation of natural selection signals for African cattle populations (AFH) breeds

The iHS (Integrated Haplotype Score) method of the HAPBIN v.1.3.0 program was used in order to discover the natural selection signal through a specific destruction on the genome. For options other than '-f 0.01', the default option was used, and the genotype of the species was analyzed considering African buffalo (AFB) as the outgroup as the genotype of the most recent ancestor.

After calculating the iHS value of each SNP, the SNPs were grouped by frequency and normalized. After that, for each 50 kb region, the proportion of SNPs having an absolute value of iHS of 2 or more among SNPs in the region was calculated. Regions with less than 10 SNPs among genetic regions were removed, and the top 1% regions with a high proportion of SNPs from the genetic regions obtained through this were determined as candidate regions showing the natural selection signal.

Experimental Example 4: Analysis of local ancestry inference of AFH varieties

The LOTER package was used to analyze the genotypes derived from taurine or indicine locally in the AFH genome. Assuming that the genetic information of AFH varieties is mixed with genetic information derived from Eurasian Taurine (EAT) and American-Australian-Asian Indicine (AAI), a total of 103 EAT varieties and 56 AAIs are used as reference groups. did Using LOTER, it was confirmed whether the genetic information of AFH belonged to taurine or indicine, and the frequency of genotypes derived from both ancestors was calculated. The ratio of each ancestor obtained through this was calculated as the average value for each 50kb genetic region, and the upper or lower 0.5% of these genetic regions were further filtered based on the Fst value (<0.2296). At this time, the remaining genetic regions were judged as regions showing taurine information or lost taurine information.

Example 1: Search for signals of natural selection through Integrated haplotype score (iHS)

The signal of natural selection called “selective sweep” was searched for using the Integrated haplotype score (iHS) method.

After dividing the genome of the AFH variety into regions (windows) with a length of 50 kb, the analysis was carried out. Among them, the regions of the top 1% with the highest proportion of SNPs with absolute iHS values greater than 2 (496 in total) were judged as candidate regions. did. (See Fig. 1). In addition, through the above analysis, a total of 494 genes overlapped with the region showing the natural selection signal were identified. It was confirmed that there is a statistical relationship with gene ontology. (FDR-adjusted P < 0.05).

The top 496 regions found through iHS analysis had, on average, 26.14% of genetic information from taurine ancestors, lower than the whole genome average (32.49%) (see Fig. 2), and the empirical distribution generated through the resampling method. ), it was confirmed that the value deviates from From these results, it was confirmed that the gene region subjected to natural selection was biased toward the region derived from indicine.

Example 2: LOTER analysis

After iHS analysis, LOTER analysis was used to determine which ancestor the genetic information of each genetic region was derived from throughout the whole genome. Through this, regions showing a high proportion of taurine or indicine genotypes as the top 0.5% were identified, and 63 regions out of a total of 496 genetic regions were identified through iHS analysis and used for subsequent analysis. After additional filtering (<0.2296) based on the Fst value indicating the level of reproductive isolation between the two groups, a total of 16 genomic regions were identified as a result of combining the regions located next to each other.

As a result of the analysis, as shown in Table 1 below, the genetic information of taurine in three of them and the genetic information of indicine in 13 were high. In addition, the genetic regions showing taurine and indicine information and showing natural selection signals were found to overlap with 9 and 51 genes, respectively.

BTA ^a Region (Mb) #Windows Proportion of SNPs with |iHS|
≥ 2 (%) Ancestry (%) F _st identified genes Regions with an excess of indicine ancestry 3 120.30-120.40 2 67.74% 93.02% 0.3390 PASK, PPP1R7, SNED1, MTERF4 3 120.45-120.55 2 63.33% 92.86% 0.2913 SEPTIN2, FARP2, HDLBP 3 120.60-120.65 One 79.35% 92.62% 0.2875 FARP2, STK25, BOK 3 120.70-120.80 2 83.36% 92.62% 0.2553 ING5, D2HGDH, THAP4, ATG4B, DTYMK 3 120.85-120.90 One 79.25% 92.62% 0.3182 RTP5 7 49.75-49.80 One 65.74% 92.62% 0.3817 KDM3B 7 50.05-50.25 4 67.90% 91.28% 0.4179
CTNNA1, LRRTM2, ENSBTAG00000004415 7 50.30-50.45 3 75.17% 91.28% 0.6321 SIL1 7 50.55-51.15 12 86.06% 92.05%
0.4861
PSD2, NRG2, DNAJC18,
ECSCR, SMIM33, STING1
CXXC5, UBE2D2, MATR3,
PAIP2, SLC23A1, MZB1,
PROB1, SPATA24 13 56.95-57.00 One 82.80% 93.58% 0.3090 - 13 57.05-57.10 One 73.94%g 93.76% 0.2685 EDN3 13 57.15-57.65 10 81.95% 92.69% 0.3114 PRELID3B, ATP5F1E, TUBB1, CTSZ, NELFCD, ZNF831, GNAS 9 39.65-39.85 4 67.07% 92.44% 0.2982 STAC2, FBXL20, MED1, PLXDC1, CACNB1, RPL19,
ENSBTAG00000008368, ENSBTAG00000050597 Regions with an excess of taurine ancestry 10 92.15-92.25 2 72.23% 59.98% 0.3211 CEP128,
ENSBTAG00000047322 11 14.40-14.45 One 67.08% 61.19% 0.4337 - 11 14.65-14.85 4 78.31% 61.34% 0.2870 MEMO1, DPY30, SPAST, SLC30A6, NLRC4,
ENSBTAG00000048521,
ENSBTAG00000049576

As can be seen in Table 1, the longest genetic region was a region of 600 kb in length and was located on the 7th chromosome. A region containing a total of 12 successive genetic regions, showing an average of 92.05% of indicine ancestor genetic information, which is much higher than the average genome value of 67.51%. In the downstream of this region, it was confirmed that the 150, 200, and 50 kb long regions showed 91.28%, 91.28%, and 92.62% of indicine ancestry information, respectively, as relatively short regions.

As can be seen in FIGS. 3A and 3B , the above four genetic regions were distributed over a total length of 1.40Mb of the 7th chromosome (49.75-51.15Mb). These genetic regions showed low genetic diversity (reduced level of diversity) in African Humped Cattle or AFH, including African indicine, Sanga, Zenga, and Sheko native varieties, and European Taurine (EUT). and Eurasian Taurine (EAT), including Asian Taurine (AST), was confirmed to show a difference in genetic information. In terms of haplotype information, it was confirmed that the genetic distance between AFH and American-Australian-Asian Indicine (AAI) was closer than between AFH and EAT.

18 genes were found in the genetic region. Among them, MATR3, MZB1, and STING1 genes were related to immune function, and DNAJC18 gene was related to heat resistance function. The PAIP2 and SPATA24 genes are reproduction-related genes and were found at the same location. The reproductive-related SEPTIN2 gene was found in the genetic region located on chromosome 3, SNP information of the heat resistance gene found in AFH in Table 2 below, and information about each gene in which SNP was found in Table 3 below.

chromosome SNP location SNP gene genetic region iHS score Ancestry 7 50671650 C>T DNAJC18 Downstream 3.46945 0.919463087 7 50677239 G>A DNAJC18 Downstream 3.20283 0.919463087 7 50678106 T>C DNAJC18 Downstream 3.73749 0.919463087 7 50693167 C>T DNAJC18 Gene 3.53833 0.919463087 7 50695486 T>C DNAJC18 Gene -3.04383 0.919463087 7 50697429 C>T DNAJC18 Gene 3.5815 0.919463087 7 50706143 C>T DNAJC18 Gene 3.64601 0.919463087 7 50709155 T>C DNAJC18 Gene -3.14025 0.919463087 7 50709950 G>C DNAJC18 Gene 3.64393 0.919463087 7 50710749 G>A DNAJC18 Gene 3.64505 0.919463087 7 50710779 T>C DNAJC18 Gene 3.64505 0.919463087 7 50713321 C>T DNAJC18 Upstream 3.6325 0.919463087 7 50713429 G>A DNAJC18 Upstream -3.12964 0.919463087 7 50714200 T>C DNAJC18 Upstream -3.13134 0.919463087 7 50714201 G>A DNAJC18 Upstream -3.13134 0.919463087 7 50714716 G>T DNAJC18 Upstream 3.63433 0.919463087 7 50715099 G>A DNAJC18 Upstream 3.61861 0.919463087 7 50716374 T>C DNAJC18 Upstream -3.11672 0.919463087 7 50716447 G>T DNAJC18 Upstream -3.11672 0.919463087 7 50716635 G>A DNAJC18 Upstream 3.61861 0.919463087 7 50716649 C>A DNAJC18 Upstream 3.74522 0.919463087 7 50716754 C>T DNAJC18 Upstream -3.11549 0.919463087 7 50716922 A>G DNAJC18 Upstream 3.61729 0.919463087 7 50717030 C>T DNAJC18 Upstream -3.11549 0.919463087 7 50718495 G>A DNAJC18 Upstream -3.12797 0.919463087 7 50719292 A>G DNAJC18 Upstream 3.63233 0.919463087 7 50719731 A>G DNAJC18 Upstream 3.59312 0.919463087 7 50720099 C>T DNAJC18 Upstream 3.59312 0.919463087 7 50720277 A>T DNAJC18 Upstream -3.09303 0.919463087 7 50720429 C>T DNAJC18 Upstream 3.59312 0.919463087

SEQ ID NO: denomination position (chromosome: sequence position) gene direction One DNAJC18_Downstream 7:50670729-50680729 - 2 DNAJC18_Gene 7:50680729-50712129 - 3 DNAJC18_Upstream 7:50712129-50722129 -

The region where Taurine ancestry information was found with the highest level (61.34%) was 200kb long and was located at 11:14.65-14.85Mb. Like the regions found on chromosome 7, it showed low genetic diversity, and it was confirmed that the genetic difference between AFH and AAI increased in that region, and that the share of haplotype information shared between EAT and AFH was also high. Confirmed. The region includes seven genes, and among them, NLRC4 was confirmed to be a gene related to the inflammatory response and tick infestation.

As described above in detail a specific part of the present invention, for those of ordinary skill in the art, this specific description is only a preferred embodiment, and it is clear that the scope of the present invention is not limited thereto.

Accordingly, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

<110> Seoul National University R&DB Foundation C&K genomics Inc. <120> Single nucleotide polymorphisms associated with heat resistance of African indicine breeds and their application <130> PN200325 <160> 3 <170> KoPatentIn 3.0 <210> 1 <211> 10001 <212> DNA <213> Artificial Sequence <220> <223> DNAJC18_Downstream <400> 1 tggggtgtct cagacactta acacagcaca tacacacata gcccttcaag tttggagtta 60 aattcacaaa gggcgctggg gtaaccaggg ctgtttatat aggaaggaat ttaataccaa 120 gagtaagagg gatagtgcct actagacaac caagtagtcc ttccctaggc tacaacccca 180 tacccccatg gtccccacta tcatccaagg gaggaaacca ggggagtaac aggggctgga 240 gactcgaggc gcatcctcag acttctgagc acaggaatgg cctggcaggt ggtggtccaa 300 aaggcaggca gggaggaaga aaggggtgtc ttgcctttta ccctgcctct tccctggggt 360 ccaggccact gtgaggcctg agggaagtac ccacagggac agggtcagga tggggacctc 420 cagagcagag aaggcagagg ttgggcttta cagcagataa caggaaggcg acaaccattt 480 tatttttccc tgggacgctg acggctccgg gcctgacgcg tccccgaggc tttcttatgc 540 ttctggtcca gcacttgggc catgtagttc tcttgctgct tctggatccg gaagtcagac 600 atgtgattcc tgcaacggag ccggctgggc gtgggctggg gacagcccag ggtagggagc 660 agaggtctgg ctccccactt ccatcaacaa cccccagcct ggcccagacc cctcacctga 720 agatctcttt ctgctggctg ataacttgct gcaactcctt aatctcattc tctttgccat 780 taagtatatc ttcttgcttt ataatgtgag actcaatttc tgtgggggag agaggcaggg 840 aggaggaggg gttcccatcc catgggctct ttcctgagat aaggcattcc ctcatgggtc 900 ttttcagaac ttggctcccc ccgacccagg ggttgtccaa gagtagttgc taatgcccgt 960 gtaaggatga gtttattata gagtcctgat ggggtggggg tgtgtgtcag tgacttcaca 1020 actctattct ggaccaggtg gctcgcctgt gtgcatgcaa aggcccttgg agagagatgg 1080 tcagccctgc cctcaactcc tgcagctctt tacagatatc acctccttgg tgagaccctt 1140 cctagtctaa aactacaacc cttcacttcc caatatttcc tatcccattt tcctgcttta 1200 tttccctttt agcacttaaa tctcacagac ctcatgtttt gctcatgtct tctcctctac 1260 taggttataa ggtccacaag gacaaagatt tttgttttat tcacagcttt aaccctaagg 1320 cctaaaacag tacctggcac atagtattaa atagatgcac aatacattta tttgtggaat 1380 gaataaacaa gatgtcccct gggagtttaa gtcttagaaa catcgcctag ttcagtcatt 1440 gcaaaccaac gcctagttca gtcattgcaa accacccacc cactgagggc tagcaggatg 1500 cccagcccta tcttagagct tcaggtgaga agacaggacc ccagggagat agtggtggca 1560 aagcttaccc ccagattaag tgcaaggatg agttttgtgt cttggcttaa aatccaatga 1620 ggctgtaggg atctggggtt gcttcattta cataatgtct ctcagggagc cctgaaaaat 1680 tctgatcctc caagaagaaa atcctaaagc atttaaaggc tagacaaaga ttaactagaa 1740 tgtaagctct gaaaatgcaa aaatgctgaa gcagttaagg tttcagaaga gaaaatctgg 1800 taaaaaatta cagcatattt tgaacaggga catgggtgaa ggaaaagaag aaaaatccgg 1860 gctcactggg tcacaggacc agtgctattt aacctccttc cccaggtcag tcagtgaatc 1920 caatctgatt gcttatgaag ttcttatggg aaaaaagcat cagaaatttg tttgaatctt 1980 aaaataaatg agagctattt agactaaatg gcacccaatg agagagtggt tagaaaagac 2040 tctcagaaga gtaaggaact atgtaagtat acaaccaact tggaaccaca taaagggagg 2100 tgtggggcgt tagaagtcag gccagggggt ggctcaggtc cccagatctg ctcaggtgag 2160 ggcttgtgcc cttctggagg aattcagaac attctgcagg gaggcagcca gctcctcccc 2220 tatgactcac acacactttc cactagcaga cgcagagccg tgaggccatg gctgatctct 2280 aggaaatgtc ctagaggaag ggcctcacca ctaggagaga agcctaggtt cctttt aggg 2340 aagtgggggg aaacccaggg aaaagggaag gactagggga gcagcatctt tgggtatgga 2400 ggcattgcct gcctgttggg ccacaggaga ggccatgagg ccatgaggtt gccttagaca 2460 accatgaggc ttgttagtcc acctggccag atcaagatgg cagcctgctg gcatctttcc 2520 agtgagagta ctcaccaggg gaaagagaag actgcattgg taggtagggg gtgcccaaga 2580 ctccaggtga gggcccatca ggagagaccc taaacctctt ttgggcttga actctgatct 2640 actcaaaatg aacactgcag tgaatgaagg aaatgttgca tatggaattc atatctactt 2700 cttaaagtag gtaaggggag tttggcattc tgggaaaatg aaaaggagtt ctaaggtctc 2760 aggacactga atagctgact gaaagtgcct cgaactcaga gcaccagctg agccagcaag 2820 tggccacaga catctgcaca gagcagaggt gaaaactaat ggcccacaga catattgtcc 2880 ttgggccata caatacttta aacatttctg aatttttaaa tttagtgcca acatgaaaaa 2940 gtcagaaaag cttccattag aatgtgcttt ccagagtctt ttttaaagat ctgactgtac 3000 tgggcccctg ttcccacagc agaggtgcta ggcccttggg aaggagtatg gctctccaca 3060 ttaccacagt ccccatttct ccctgttggg tccccagcac tcagactaag gatcagcttc 3120 catacaggac tattctttcc ctgaaaacag agttatgagt aaagtacact atttctatta 3 180 gtcaccaaaa gtgggaaaac aaaaaggtgg actaagaagg tcgtatgttt caagaaaaat 3240 gggagcgagc ccgttccttt atggccatga agaatattgt acaccaacca cttccctcat 3300 ttccgtggcc tgcctggtcc ctgaaggcct ttgcgaaggt gtggcccgct tgggctggcc 3360 tgtatcctca gccggggaca aaagttgggg tgaaaccctg ccctggacac aggcactggg 3420 ctgttaggga agtgcccatg taagcaagcg tgggagggag tgcgggcctc ccgcctacca 3480 ttgcacttgg tgatgagctg gtccttcttg ctggactcct gcagggctct gctcttgaca 3540 ctggagagcc tgagtgaaga agataaggaa ggtgggaagg ctgatgggca gatgggaccc 3600 ccaactcttc ccccactacc aaggtccact cacgcctttt caaaggccag cttctctttc 3660 tccagctgtt tagacagcac ctctacctct ccgagggcaa actgcaactt ctcctcttcc 3720 ttggccagga gaaccttggt cttggcatgg gctgctttct cttcctgcag gggcagcagc 3780 agtgacctta tcaccacaag ccacgctgag tctcctaggc ccccttaatc agcccccttt 3840 ctcagacagg catgtagaag gcatggtaca ggaagattct gtcctcacta tgggggctgg 3900 aggtgctcag agaccagtgc tcaaggactg cccaaccact aggatcctgg gaaatggggg 3960 aagatgagaa gctgacagag gccattactt accaccagct ttttctccac tgcctggaac 4020 tc ttctttac tgacaaaatt ttcatcctgg agaatcatct gcaacagagc ctggttcagg 4080 gagggcctgg ggttgtggtg ggaggggttt ctctaggcac cagggcctgg gatagataat 4140 ggatgatgcc cagggcatgt acgacaacac tggggcccaa ggaagggaga gttcattcca 4200 caagattcac tgatgctcaa ctctaggtcc tgcacacaag gaataagtct catggagaag 4260 acaccatggc caccatgatg tgcgctatga ggcctttgcc tcagcctggg gaagggtcag 4320 aacttcttca agggaaggga tgtcaccctg atatctgact gctaaatgtg ggtgggccag 4380 aaaaaaggat ggaaaagaga agtgttccag ataatgagaa cacacatgta aagaccaggt 4440 gtcaacagaa aggcctgggt acctttaggt aattcagtgt tcagagcggc attcaaacat 4500 tggagtgaga aatagatgtg aagtcagttt tatctatgtc ctcaccttct tatcaagata 4560 aaaaaaacag aaaaatcata tatcccactt ggccatgatt ttaccataca tgttcaggaa 4620 atttgtggct ccagtgcttt gtcccagctt tgcatttgat tgacaaggca aaagcaaagg 4680 acataaaggg gttggaagcc cttaagtcaa catgtgacga gcttacagca ctcagcatgg 4740 agagtgggtt ccacaatagg gatgatgtta tagttactct gtacacagtg aagtcatctc 4800 caattacaaa tggggaaact gaagctcagt gaataggctg ccatatctac caggccctgg 4860 acaaccag gc aaagagagca gggaacaact tggttttctc taaaatttct gggtaaatgt 4920 tcctgagaca tagtcccttt tgggctctac ctcgctacca gccacaacac atcggggccc 4980 cgcctcctcc ggatgacgcc attggcaatg tggtctcggc aaggccctgg catagggaca 5040 gggcctggcc ccgcccccac cacgtaggcg tccagtcagg cctcccccag ctctctggct 5100 gcgtaccacg ttcctcagct ggtggatcag ttcctcttga gactcaatca cgtcccgcag 5160 ttgatcaaag gcgagacaca ccgaccctga ccccccctgc gaccacccga cgggcgtcgc 5220 catctttcgc ccccgccagc tcgttggaaa tggctgccgc gggctgcttc accgctgacg 5280 cccaccaatg agaatgcggc gccacaaagc accctggcaa ccaggggcgg ggacttcctt 5340 tgtttgcagg gatggggccc gcctcttaaa agagtcgctc tcttacagga gccgcctcgg 5400 gcagagaacc tgcagcaccg ttagtaattt aggtttatag ggaatttgca cgtttgttat 5460 gtggtgtctt tattacgcag tcctgaggag cgggcagggc ccggtttaaa cttactctaa 5520 gggctgagga gcaggcaaca ttcacaccgg cccacagcct tcccccagga cccattaaac 5580 tacatgcaag accttgaagc aagagtcatg gacaacccct gcgtgtgtct ttatcccact 5640 ctaaactctg aatttcccgc atagtatgag actaatgtga tccctcttcg aagctcttag 5700 gaacacactt agt acttagg gggcctgagc ttcccaatga agggaagcta aaaaatatta 5760 aagaaaagtg acttaatcta agataagagt ccttatattc tgggaccagg ctttgggcaa 5820 cctgaagagt ctgaagatat tagaactgaa gtcaggcttt gaaggagact tgtaaaatgg 5880 aggaaaggaa aaataaagtg attacatatc cccaccaatc cccgatgagg gtttgttttg 5940 aaacccaaat aagattatgc ttggggtaaa ccagaactaa tatttatcaa gtattcacta 6000 tttcatgata ctgttatgcc ttattagtat tatttcatgt gaccttcata ataaccttgt 6060 gaactcgatt ttaatatccc cattttacag atgagaaaat gtagtttcta tgagaatgaa 6120 taacccagta agtcaaagtc acccattgag ctttgaaaca gtgttttgac gggggcagtt 6180 tgatttcaga gtccatgttc ttaatcatga tgctatgtag tattttgtga acataatgga 6240 ttctgaagtt atggaatttg aaactcccaa gtaggaaatg actatattaa gggatttggg 6300 aaaatatgtt gtcaatgacc acaaagggaa gttgaaatgg caggtaagtc acttagggtg 6360 ctctcctcag ctgcctgggt atctcttttt ggctggatgt tgtgacctga caaaacatga 6420 tcattcctgt ggatctccca gtggtaatcc ttgttttatt tttccctttt tttctttcct 6480 tttttattgt gtttcttctg atgtccttga gggtaaggcc acctctgtgg atcaatctct 6540 aaggtcatag atggacctt t ctgtgatgga aacgttctgc actgtccagt gccagcagcc 6600 actagcctca cgtggcttgg cagcacttga aatatggctc atgtgactga ggagtgcttt 6660 ttaaatattt aataaaaatt ctaattagtt aacattgaca ttcaaatagc catatgtggc 6720 tagtggctat ggtattggat tatcatgggc ctgtaggatg ttctgtgatt tatgtacttc 6780 aggacagagc cctcttggaa ctttgtgttt caaatgcgaa tgttcctctg aatcaccctg 6840 gacaaagctg tacattgctc tggggcagcg atccctgagt ttctcctctc tagggtcttt 6900 catgaggtga tgctttatat cccttgccac tcaaagatca ctttcaccag ctgctctcca 6960 tgttcagggc aaaactctgg agtgaaatag gctcccgcag ccagttccta gggggtggga 7020 agttgtgagc aagagagaac cctggatcca ggcccacggc atactagcac ggacttgatc 7080 tgggatggtc cctctgtcca gagactgcca agactcccac tctgtgacaa cagacacttc 7140 aggttcttcg ccagctgggg ctcagcaagg ctcaggataa catgacatca cagatagctt 7200 tgtgatcctg ctccaggaag gactgccttt cttcacaggg ctgtcatcgt ggttgatgtt 7260 ctggttatct attgctctgt taacgaacca cccaaaatta atagcttcaa aaaaagtcat 7320 tttgctcaga aatcttcagc ttgggcaaag ttcaatggtg aaatcttatt tctgctccat 7380 gaggaccagg gttcttggtg cctc tccacg ggctgtttgg gctctcttat agcatgtgac 7440 tgcatccaaa gtgacaaaac attctggtgg tccagtagtt agttaggacc ctgtgcttcc 7500 actgctggtg acctgggttc gatccctagt cagggaacta agatcctgca agctgagtct 7560 gcggccaata aaccctcaaa atgacaaaat gagagatgcg tttttcttaa gctctgggcc 7620 taaaaactgg gttagtgtca cttctgccca actctattgg ttaaagaaac cacagagtac 7680 agattcaagc agaagggaca cctcctacct tttgatggga gtagtttcag agaatttgag 7740 ctgcacttcc agaggaagtc ctgacaggtg tcatgtcatt tccttccaag gttcagttca 7800 gataaaacca gactggaact tagtctaatg ggggagacag ataataatca aataaacacc 7860 caaataaatc ccaatttaag tcattattca agcaagaaaa agaggaataa gagtaattta 7920 aaaggagatt tagcccagcc tgggtgtcag gaaaggctgt aaggaaaaga tgtttgagct 7980 gaatgaaacc tattaaggta accaggaagc acttaacaaa ggtgggcagg ggcagggaag 8040 tgggaaaggg taccagaaag agggaaatgt gaatacaatg tcccttgggg aagaaggttg 8100 gcagagtgta gtaactaaaa ggttagtatg gcagaaaaga aaacaatata caggacatca 8160 tgctaggata gaggccagaa acttggtaga ggcttgttaa gcattaggcc atgcagaaga 8220 ctttactgat tcttattatg agagcaaaag aaagctactg aaatatttta agcaagaggg 8280 tgttatgctg taatttatgc tttgaaacta tgtctctggt tgcagcaagg tgtacagatt 8340 ggaggggacc ctgagtggat gcacagagac tacttatact taggcaacta ctgcagtggt 8400 ttctgtcaag gtggatggta gtttgggagt cagatggtat cagaggagac agagtagatg 8460 gatttggggt gtcctggagg taaaactgat aggacttaac aatggaaggg atatggggaa 8520 agggagccga ggtgtttggt tttttactgc tgctgctgct gctaagttgc ttcagtcgtg 8580 tccgactctg tgcgacccca tagatggcag cccaccaggc tcccccatcc ctgggattct 8640 ccaggcaaga acactggagt gggttgccat ttccttctcc aatgcaggaa agtgaaaagt 8700 gaaagtgaag ttgctcagtc gtgtccgact cttagtgatc ccatggactg cagcccacca 8760 ggctcctccg tccataggat tttccaggca agagtactgg agtggggtgc catcgccttc 8820 tctgttctac tgctgcagac taccacaaaa tttaatggct taaaacagta cctagtgtat 8880 tacttgctca agattctgca gtttgaactg ggcttggtat gcagctcatt tctgctgtga 8940 tcatcagcag gggtggcata accagtttgg ctcagatgat acgtgtctgg gttcagttct 9000 tgctgccagc tcagttgttt ggttctcctc tggatggctt ctccacgcag ttagcttagt 9060 ggtctcaggg tggtaggacc taaatggaag ctgct ggttt tgccacattc tattggttaa 9120 agatcaaggc cagcccaggt cagaaagggg actacacaag gtcatgaata ttaggcgtga 9180 tccattggag gataccaaat ttaaaagtct accacacaga gtcttctggg tttctgactt 9240 ttttcttgga tggatggtgg tgctattcat ggaaatggga gcatttcaat ggtgttgctt 9300 ccatccccat ttttaatcac tgcactctgt aagaaggatg attacatttc aagtctcttt 9360 tccctcagaa ttgttttcat tgatccttca tcctggatta agtcccattg ttatagattc 9420 ctttggttcc ccctttgaca ttcatcccac tgggaactat tgtctaattg gcctccctgt 9480 tagatgagag ctccaggagc agtggtgctg gctttcactg tttcctgatc tccatcccaa 9540 aagctcagct cacctggcac atagtaagtt ctccatcttt gctgaattga gtaatcttaa 9600 tctttcttca catgaaaggt attatgatta tattaacaat gtcctgtatt tctaagccat 9660 gactaaccta acaggaccat gaggaacaaa accagcaagg aactgggata acacctgatg 9720 ttcagatgct aagtggattt gatgtatctg ttggttgtaa taagctgcag ctgctgatat 9780 ccgatggaaa acaaatattg aattggtggt gtcaccgatg ctatctattg ttctaaacac 9840 caataattcc attgatgtat gagggaacac tggagcagag gtcaagagct ctggtctagt 9900 ctagtcctaa gtggctccaa ggagtcaggc aatatgcagg acctaagagt gccatgggca 9960 aaggtaaaag aacaaaggaa aactataaag ccattctttt a 10001 <210> 2 <211> 31401 <212> DNA <213> Artificial Sequence <220> <223> DNAJC18_Gene <400> 2 aatataaaat catttaattt ctttctctgt taaaatatac tcagtccaga tgctgtggat 60 caggtgacca gatttttcag gattctcact ccatcataag ctgattccat ctgtgtcaaa 120 cctggttctg gtcacagaac attcacatct ttaattacaa atactcaatg tgatgcaaat 180 catgtaatca tatctaacac ataggtatat agtttctcgg tggtcactgt acaaggtatc 240 aacaatgcag gctcaaaaaa taagatgtat agtaagttgg tcctttccaa aaatatatta 300 ggacactaag cttacacctt aaaggaataa aaaataaata ctaattaagc gctagctcct 360 gtgaggaaca gtgcatgcag acagtgaact cacatgtgtg gccaattcat aaaagaaagc 420 tctggaatcc caacagtgtc tcctcataaa gccagtcctg agtgtgagga cataaatgag 480 aataacacat agcttggcca ggctgctgga ttccacgtgg ggtttggagt tcagcataat 540 ttctaacagt ttccaacaga aagaacagca atggtgtttt cttactggtc acttttccca 600 aactgtgagg cctggcgctg atcaaggaag ttcagaagcg ttctagtttg agcagatgaa 660 ccccagagt c tacagat gcct tcagaggt c tacagat gcct ga 720 gaagggtttt tatcccaccc tcctcactgc tgatcatgtg ctgcagaagg cacttccatt 780 cttttgagag cttcggtttc tgcttgagcc tcaagcccta cttcttagcc ccatcgccag 840 tctgcattca tgctcacaaa aatagtcgag agctgcatgc caagttctga cagcaagaat 900 aagccataac tagaaagttt atatatgata aaaccttcca aatgatgtag gcaagaggaa 960 aacacttatt ttaccaccat gcaaacaacc catctaaaaa tgcaacccag gaaattatag 1020 tttaacaata ttctggacag taccaaatag attaaactat cctccaatcc aaagcacata 1080 ctatcccctt gaggaatgtg atgtgagtgt tgtcgtctat taggtaataa ataaaacggt 1140 accatcagtg ctataaaaac agacaataga cctgacatct aatgaagtga gttagtattg 1200 cctctattta gtatcatcac agataactca tctctgccat gaaatccata aaatgtgtga 1260 ctacactatt catgggcacc tagagcaggg acttttaacc tgtcaagtca gtttaattat 1320 ggtccatttt ctagcagagt ttggggttgg ggagagtaga gctttgattt ttgtatatat 1380 tggaaaacaa ccttctacag cccatctggc tcacaggcac attttaccat gtaaatccct 1440 tcattcactc caaggggaac aaggaagtcc ccccaaaccc agaagataga cctagttggg 1500 tgacaaaaat ttggccatct ctggagctgc tctccctccc tggaactgag caggtgtccc 1560 cta ccagcca cgaaagcccc actctttacc tctgcactca tctcctgacc ctgtgggatg 1620 ctcccaggtg aaccatacta aatacgtctg cttaaatcat ggctctgcca ctgaaaggag 1680 aaactctact ttggaaatga gttttaatgt agactctcct tgcaacagga acttttcttc 1740 catcctgaaa acacatggag tgttgactac agtcacctac tgatccttct aggttcagat 1800 tcatgataga atctgggcag ctcaatcctg tagctcacct ccacagaaag tcccagaacc 1860 acacagtcca tgcgatggct tccctcttcc ttgtcaccag tgggaagagt tgcctctttt 1920 tgtttggtat ggtaagtggt gtagtgaggt gtgctgagtg gatgcaggac tgccatagaa 1980 aggataccag gttccaggct gggaggtggt gcaggagggg catccttggg aaagcttctg 2040 cttttactaa tatggctaga atcacctgag caagggggat gttcactcac tctgtacttg 2100 ataatcagcc taccatctag ctctgccaca accagttgtg tgacccaggt caaccacgta 2160 gcctctttgg gctgatggtc ttatccatac caggcactta acagacattg actacatgaa 2220 aaaagcatct cctatcacaa ccacaggtca ctgggttaaa caaagtgact gctgggccaa 2280 caccagatcc caccaggccc cagagccaca gactaagggt ctttcctatc tatctctagg 2340 tccgagcagc ctggctgggt gggaatgcct cctcttaggt ctgaccactg tacctctatg 2400 gggtatagg g actcagctca agtgtgatgc tgcaaggaaa aatgacaaag aacattctct 2460 gtgttctata atggactagg gaaactgcac tgaacagctg agactcagac ttgcttttta 2520 ccttcacccc tggcccctgt ttctgtgtgt ccactgaact gtttactcag taactcttaa 2580 tttctgggcc ctttgtaaga gagagggtta agaaagtaaa agaaactgaa atcaggtaat 2640 ttgctaatgc cagtaactaa acagagatta ggcaaatgga aaactgtgga caacagtttg 2700 cgtttctctg tattcccctt tgaaaatgga aagctgagtt gttacaacaa ctgcatctgg 2760 tggggagatg actctaacaa caggagaggt ataatttctc atggcacatt gtaaacaacc 2820 aatattaagc tgctttggct aagcctagga atgactggat attgagattt cactgaagaa 2880 ggaagaaaat atgaatctgc tctgggatgc cctgaactac attcctagag gcaggtaggt 2940 acggctcagg gggctggatc taatgcttgg ccgtgaattt agatgacctg ccccttgttc 3000 ctgaagccaa cccccaggaa ggattctgtg atgacaagtc tggctccatg ctgaagcgcc 3060 tccaactctg cccaccaaac aaagttagca aattgtgaaa tgttcatcat ttcctaattt 3120 tgtattataa aatggaatta ggaacataaa cagtaacatg tagcaaaacc ccgagcctgc 3180 gtaggactat cagcctctca gccacctctg cgtaggccaa taagtttgga aagtttttca 3240 cagttttcaa gttt cagaga ttctgctttc tgtttcagtc gttcatctct gtataatcct 3300 gccaaattgg tcaactccga ctctgaaaga taaaagaggt aaccaattca tcaagagagt 3360 tgagctcctt tgtttcactt caattaatgctt gga tttctttcaa ttaagtttaa gaaaaaaatt catttactcg agaaacatgt atcaggaagc 3480 tataccacac caggtgctga ggattaaggg acacctgtga tcctggagtc cgacagtatc 3540 tgtgctccat tacaaaagca ccggacttag ttcttatcaa tagaacattt ggcacaagac 3600 ttagggaacc tgcacgtgtg cagtaagagg agactctgtg tcaaggcttt cctacgaatg 3660 gtattcacat ggcagttggg atagaatgga gtccgatcgc ctgttcctgt catctgaaac 3720 aaaagcagct gcttcaccat tcccgagttg ctggagcata actcacacgc gctgatggac 3780 acagagccct cctctggggg cctgcctgct ctaggtgccc aggctgctgg ctgcatcctt 3840 catgccctca ccacatggga ggcccctaac tcaaagggca aaggcagctg gattctttta 3900 aaagtgaaat catctggatc tggtgacctc tatctccttg acaaaatgta tttcattttc 3960 tcctcatagt ggagaaggtc attctatttt taaccagaat ataatctctg ctgtcctccc 4020 agactcagca gcggtttggc tacctctctt tttcattctt ttgtgtagtt tcagagacac 4080 tctgaggaag gaatgcctga agagaacctc cagagacatg ttgtgtatta acaggaaggc 4140 atgcttggag ggcctgtgta gcttcaacag caggtagcta ctcagtgttt aaaggcccat 4200 acttgtgaaa atgctatttt aatcattctt aaatatgcct attcattctt aatcatcttg 4260 ctgaca gatt ttgtttttct tggtgaacac gaggcacctg tctgattaat cagcaaactg 4320 actggaaaca ggacttaata cttagccatg gggaacaatg ggcattggtt tctgaataag 4380 tctctatcga cacaaaatac tggcatttct gctcacaatg attctgttta caccaagtaa 4440 ggcccaaagt gcagattaag tgcaggactt tagggcgagg ggtggggatg catgaaaacc 4500 caccctcttc aataggggct cagagcccag gccctggccc agcagttgct tggcaagcag 4560 caagtctcct aggctccaga acaatgtgct ttcacctcgt ggtgtggggc ctgcattgcg 4620 tgccagccag agcttgcctt ggcatgggca ttctcatggg tgatgcagct ttgtggagga 4680 tcatgggccg cacccaaatg agagtccttg ctgccttgtt ccacccagag ctgggcactg 4740 actccatttg tttggctgct tcggcatgtg ggaaacctgt tcaagagggc tctgtgatcc 4800 tctggagtca cagctgcacc agtatgtaga gcagattctg tggctcagct tttccttgaa 4860 atgatttcct tagggttttt tgttactgac cttatccaga tgtctcactg cctgaaatct 4920 cattccagaa aacattaacg gctagtgatt ctttctattt tctacactga caaccactgt 4980 aataagctga tgaaagaaaa catatgtgca catatatgaa tgctttctca aaacatgctc 5040 caaaggttta gaaccctgct ccagagcaat gaccctgcag aagaacagtg tttcaaggaa 5100 caatgcccca c acccccatg atcatggttg gagaaacttc tctccagata gccactgaat 5160 ccttttgtac ttcaatgaac aaaagctcaa ctcaatgaat atgttacttt tgacaaattc 5220 tgttaatatt cagtaacttc taaggaaaaa caaaatcttt ggcaggggca gaatccaagg 5280 aacactgttt tcttctctgc ataatgtttc ccctgctcca aatgccagtg taatagatga 5340 gcagagcact gagcctttag agtaaactga cttgggcaaa ttgccaagag aagcaagtcc 5400 cagagcctcc tctcccaaaa tgctctgccc aaaagcccag atgggagtcc tgacagatcc 5460 caaccaccct catgatccag ccctgtgaga cagagctaca tctagttgag caaggccaag 5520 cgaggtctgg attctgttag gatccgggca cttactatat gctagactct atgctaagtg 5580 ttatactcat gtcatcttct ataatccata caacctcctt gggaggaagt tcattttata 5640 cccatttcaa agcctaagct tctgccctct ctgctatcct gctccagagc ttggacagtt 5700 ctcctctgcc aggcagggct ggggtctcag tcccactcag ccaggtcagg aggcagtcct 5760 gccttttatt ttagtctgtc ccagcccagg gccatggcca cctcccctag cgttggcttt 5820 attatccaga tgcagctgtg aaggggtaga cagggcgcca gtgtttacag taaaggagta 5880 ctcatagaaa ttcactctga tgaacattct aaggaagagc agagcacaga aattaacaaa 5940 agatgttaaa agacaat ggc taacctttgc taaaagcagt ttccattcgg tattttgcca 6000 agactgaact ttgttctcaa gcaggtttat ttttacaacc tccatccatg tgctaaattt 6060 aaccatgtcg gctgcacttg ctgctggatc cggagggcag atgctgcctg ttctcaaatg 6120 gcacttgggt tgtgtggcgg ctccggacag ccaccatcct taaatgccag aaatagattt 6180 ttaaggggtg tgtgctgata acccccggtt tggaacttgt gggtacacat gtcatgctca 6240 cccacttgat tatacctgga aggtgccaaa gccctggaag caggcctcac gaggatgggg 6300 acacaaggct agaagccacc ttaggtgtaa tcctttccat gaaggtgtta gtcgctgagt 6360 tgtgtccgat tctttgtgac cccatggaac gtagcctgcc gggctccaat gtccatggga 6420 ttctcctggc aagaatactg gagtgggttg ccatttcctt ctccagggga ttcttcctga 6480 cccagggatc gaaactgggt ctcctgaatt gcaggtagag tctttaccat ctgagccacc 6540 aggaaatcct aaccatgagc tctttgcata gttctagctg gtgtgtgtgt gtgtggcggg 6600 gtagggttta agaaatatat attttcacca atttattgca taaagcaaca aattcaagac 6660 cctctcatgg cctaaaacaa tggagattct taggacatac actgaccctc acctgatttg 6720 gatcaggtga tcaattataa tgcctttaag tctaaggcct ctatcttgct ctaaaattat 6780 ttcctggttt ttgaaactag aa aaacctag tttgacagtg gggtagggtt acttccagac 6840 cccttgtttt ttgttttttc aataaaaaaa gcaatcgcct cctcctgctg aattccccct 6900 tacatccaag tcaatgccca ccatgctcct aaggcactga gaggtcaagt gggctccacc 6960 ctacaggagt agacagtgga cacaaggtcc ctcagggcca gggttaggac actagccctg 7020 ctcagtgatc cctcagcagt ttcacttttt ccagtcccga gaatccactc ttcaaaagac 7080 atttgttgag catctgttct gaagcaaaga ctgccaaggg cttgggacac aaaattagta 7140 agacatggtt cttgacctca gagagcccag agtccagcag ggagactgtc atatacctag 7200 aggacaggaa cactgtgagg gtggtacttc agtaggatga gtgaagggct ttaaacacac 7260 agaggaagga gagattaact tggggtgggg ttgggatgca gatgggtcag caagcgctta 7320 gtagcactgt gactgaggat ctaaatttat cagacactgt tagatgttta ataatacccg 7380 gtgataagtt ttttaaaatg agttagtttg ctttttaaaa attaaccaaa gccaagtttt 7440 ataagataca attactaatg cctctgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg 7500 tgtgtgtgcg ggcagggggg aattcaagtg ggaagaaagg gagcctgttg gctgaggcag 7560 ggctgccttg ttacagcact ccccatgaat gctcactgca gtcgtaagcc aaatcgtaag 7620 ccaatactgt caccctcgct gtctccag ac agggaaacgg tgtggaaggc ctgttccagt 7680 aagagagtca cttttgttct gtgtggccac agctgattgg aggggaagag aaggggcacc 7740 agactcaata acaacaacct atcaaacttt ggttaggctg gagcagattt ccctctggga 7800 aatgtggaat taagaaacca tggaccaaag cagcgagcta cttaaagagg aaccttgtga 7860 cgtagaaagg ggccaatgac tggtggggtt ggggaagttg ggggaacgaa taagagggtg 7920 gcaaccaaaa taaactaggg caaatcaagg ctgtcaagga gagcttgaaa aaacgtagga 7980 aaacagttgg caaagaaagg acagaaggca gataggcaga cagatgtagc aactcagaga 8040 cagacagaca ggcaactagt cctgcctaag tccctgaaga tttcaagccc tggccacacg 8100 tatcctgtca taagcctccc tttttatcac aatatttcta atatcctgaa tggggctctg 8160 tcctgtccgt gttcacataa acatgagtag ctcagaaaga gaggagagat cgctacgctg 8220 gggatacctc agggtggcac cagctaaagc cccttttaag catcttactt tgttgtttct 8280 ccttccaaca actggtctgg atgtaatcaa tgtaatcctt ctctatcgtc ttctccaggt 8340 cacgcagaga tgcacctctg taggccttgt caaagttttt atccacgaag taaggcacct 8400 gtaggttctg ggtttctcta gaaatggtgt ggcccaaggt cctaaaacaa ggagaggctc 8460 aggtgtgctt gttatatttg gaaatatcac tct tagcact ggcctcgctg atatcatcat 8520 catactcacc tcttcataga tgcaaatacc atttctaaca catcaacatt tattatttcc 8580 caagcaaagc agtaagtgca atgttgtcat tgtcgctact ctatgagcat gttcaagtgg 8640 attcagtctc agcatattta ctttattctc ttaaatgagt catctccctc tcttctttct 8700 ccctctctct ctctttaaat ttaagacata catcatgatg ggtttacata tattgtgaaa 8760 tgattaccac aatgtctcac atgtctaata ggtaatccgc taacatctgt cttctcatat 8820 agatacaata aaaacaaaat aaagaagacc aaaaaaaaaa aaaaaaccag acaagaataa 8880 aggaaaaaat tttccttgtg atgagaactc ttactactta gtctctttaa cttttctatc 8940 atttcctatc atacagcagt gttaactact ataacactgt attgtatgtg acatacaccg 9000 tatttactaa catttatctt acaactatta gtttgcatct ttgaccatct tccttcaatt 9060 cccctccctc tctcttccac ctttggtaac cagaaatttg atctcctttt ctgagttgtt 9120 tttttttaga ttccacatat aagtgagatt atacagtatt tgtctttctc tgacttactt 9180 cgtttagcat aatgccttca gagtccctca gtgtagtcag aaatgctaga attttcattt 9240 tttagggctg aataatattc ctgtgtgtgt atatacgtgt gcgtgtgtgt gtgtaggctt 9300 tctccacgtc atggctacca taaacattgt tgctatgaa c atgaaggtgc agctacctgg 9360 attcacctct cttagactgt tttctctgtt aaaagtagaa tgctgaaagt ttaaaaagca 9420 atttatatat tctgtaggac ctggcatggc agctggcaca gagtggctac tggtaaatgt 9480 tggttgaatg aaatgattgc tacagtaaac tattcacaaa taacgagaca gcttatacca 9540 acttagttcc aatttttagg acatgataaa aagtacttca cacatagctg ttagatgaga 9600 actgcaaaaa actgagacaa tagatttcaa ttatacctcc aggcgaactc ttagtgatat 9660 ccaaaagtct ctgatgttca gaaactttaa caggtcgtgg gttaactaaa gtctaaccat 9720 aagaaagtac agaagggagc tcaaattcca gttaaaacaa tgtgatggta ctgtgcaaat 9780 ccaagctgac tttgttcaga ttgaacaaag tctatcttaa aggagttata tttgaccttt 9840 tcctagcttt tcctttgatc tcttaactca tggatcttgc tggatgtcta agagctcagg 9900 ccccgattgc tcttgcctct ttctggaaag ctgagggcag gttcctgcct acctcactgg 9960 cctgacaaga aaggaggtgg cagtagcata tggagaaata tgggaagtca ggaaatcaga 10020 gggactcagg ccagcgtgac ctgatagcaa taacctctct ctgagtctca gctttcctct 10080 ctgtaaaatg aagatattga tctagaatga tagccttaag gctatagtga tcaccaagag 10140 ctcttcctct taggtactgt tccacgacgg agtccacatt t gaaagtttt ttgtctgtta 10200 aactctaatt gtgaataatg gctcatttcc cattattaat taaaaaaaaa ccagtgattc 10260 tcaaattttg catggacaag aattatctgg aaggcttgtt aaaactcata ttgctggttc 10320 tcacctctag agtttctgat tcaatagata cagagtaagg cctaagagtt tgcacttcta 10380 acatgttgcc acttaatgct tggtctaggg atcaaacttt gagaactgat ttaactgaaa 10440 tacttaattg aagaaattta aaaattccta ccagagctca tatttttcat gcaatacaaa 10500 ctttataatg atttctatat ttttagcaaa agcaaagaaa tgagatacta tggtttgact 10560 gcacagcctt atgaataaaa gtcctaatct tgcagtttct taaattgggc ataatcacta 10620 gatcttagtt actatgttct cagattcaca ggggtgcagc gatactttca caagctcatt 10680 acggtccttt ctaactctaa catggaatgt taaagattta atctgaacct ggagttagtc 10740 tgttaatccc tcagaaacca tctgttgctg ttccgcagtt gaccagatga tgtcgctata 10800 gaacaaaaca atccaggtcc tgaatacatg ggaaactgcc tgaagtaatt ttgagtctct 10860 gcattattac tatccagctg ctcctttaaa cacaaaacca gtgttttcca caacaaaagg 10920 tgacatgaaa tgttagaaat gtaccagctt gttgttaaga aagattttaa gtatttgact 10980 tacgatttat agaacagact atatggggga ttag cagcta gcagctgagt aatcacagat 11040 atgatcacaa tcaccagaac tggaagtaac tgaataaatg cagaatatgt agtctgaaaa 11100 agaaaaaata tttggtgtaa gtactcttca tcaatataaa gcttactgaa aataggacag 11160 ttttcccaaa gaaaatatgc attactaatg cattgcacat tgcccatagt ttttctgaga 11220 aggttgtatt ttctgctgag aagtaccaaa cagctccatt ctgtctgctg aaaaggctga 11280 agggggccca cccacaacct ggactgacac tcagtaatcc tgcagcctcc ttttctaaga 11340 ccaatgaagc agacacctgt gtgctagtct gcccagcctg taccctcaca aggtcatgtt 11400 atcatgcagt tctgttgcct tggcctcagc tgagcagact gggggacatc tagcagacat 11460 ctaaccatag cagggatgat cacattcttc ctaagaatct gggattgaga ttcatcacca 11520 ttcactgcta ggcagtcact tgtaaggaca cacactcaga ggcactgtag actgaacact 11580 tgcaagggga agtagaaaag ccaggctgct tagagagaac aaagcagatg tagagagtca 11640 ggaaaactgg agacagtctg gccccaagag tcacagagag cacagctgcg tgatgcctga 11700 cagcacccca gttcctggct cccatctgtc acaaggccca gcgtgcctcc tgcccttgag 11760 cttcatgaga ctccctcgat ctcttaattt aaaccccctt tagtactgaa tattataaag 11820 gcggtcccta ttctctgcag tcaagaa aat ctttttcaag acagccccat aggggaacag 11880 atttcccggg atagtcccca aagaatgggt gggggagggg gtaccattct ggcctatttg 11940 ggccagttat ccagctcttc ttaccatgct tctgctttcc taaagcacgc tttagcacat 12000 atctcaagct gaacttcaaa aacctgatct tactccataa gggttagttt cagtatgagc 12060 gtttatgggc cgctattgtc ctatcctccc caggtacctg aggcttatct tcttcctctt 12120 ccttccgtgt ctgcattctc tcatgtcggt gccgccggcg gtaatagtgg gtgtcatctg 12180 tcacatttga aaacatatga atatttcctg gaaaagaaag aaaaatactt agcacgcaaa 12240 tatggcacac acctcttcct tgccttttcc aaggtccgtg cttccaacta ggttctttga 12300 gggtagggta ttgtcccatt ctacccaata gaaaaataag acagtaggta tatcaagtga 12360 accagctgaa aatgggacat gcttagctct ctcaccctta tttaaacagt cctttgtaac 12420 tagcgagtga gagcaacctg agaggcagag aaagaaacag cacttgggcc atgaatcctg 12480 tctgacatgc tcttatctgc cttcttgccc aatatcagat tatgtcacta gtatctttaa 12540 ttgcaactcc acaaaattca atctacttcc aaggcattta attaacccaa cttccaattc 12600 tctgggggta tttttctaac ctttaccttt atataagtgc tagtttacaa taattataat 12660 ataggaattt tgcttctaat ttataaaaca cttaaaaaat attggttggt attatcagcc 12720 tcattttttg gtgaggaaac aagatcagac ggactaagta aaaatcatgc atagttggag 12780 gctggagacg acttcaacct cagcttatct gattccaaag tcttgtttcc accataccac 12840 ggcccctcta ttcagatcct gaaaatatat atgactgttt ttggccacac cacatggcat 12900 atgggatctt agctctccga accgggatca aacctgagcc ccctgcagta gaagtgcaga 12960 gtcttaacca ctggaccacc agggaagttc ctaagacttc ttaaatgaaa aaagttggga 13020 tactaacctg taggaaaatg tcctccaaag aagacattga acagctcttc tggagtgatg 13080 tcggtttcaa agtccctgta ataattataa ggtctggctc gaggggcagt gaaagtcacc 13140 tgttcatctc catattcatc atagcggagt ctcttgtcag gattgctcag gactgcaaag 13200 gcatttccta ttgctgcaac aaaaacaaaa agcaccagag tacacagtct gttgcggctg 13260 agggtaacca cagtagtagc agtgaagagt tggtgctgga caacttcaca gtgacccaag 13320 gagaggtgct tgggcctgga aagttcaccg aagtgcttgc ccttgaaatc ttcccaagat 13380 ctagtttgat aagccttaaa cactggagtc aagataaact tagaagtact gtgctccttc 13440 atcaaaaatg tgttatttca cttgagagca atagtttatg gtcagaaaca gtttatacat 13500 aagcaatggt at tctaacat gaagtcacat acagtctcct gaataaaata ataaataaaa 13560 atcctcctac ttggttaaaa gcaaagtcct cagtcatcag tcttatttca gcttactttt 13620 aaacatagtc tttagaaatt aggaaatgta tctccaggga attccctggt cgtccagtgg 13680 ttagaacctg tgcttccact accgagggtc caggtttatt ccctggttgg ggaactaaga 13740 tcccacaagc cagtcattgt ggcttaaaaa aacaaaaaaa atgtccagtg cctcagtgag 13800 ggatcaaagt ctgaatagac tacataagca taccatcact tctccctgtg cacaaatgtg 13860 caaactttgt ccaatttcat ttaaacatac agtcagtacc accaatcaaa gaatcaccaa 13920 agaaatctta aaaaaaaaaa tcttttaagt cattcccacc tgcctacctt taattcagtc 13980 aacactatca gaaacatagt cagaaagagc atttccttgt gagtcttcta gtcagggaac 14040 tgtattaagt atttatagca atagaacagt taagcagaaa aaaaaaaaac cagaaaaacc 14100 taaacagagt tgtttaaaca ttcacatccc ttgaaatctg gcagatttct tagagaaatg 14160 gcatcagtct gtgacgtacc ctacatgaag gtatgaggag aatgtggttt atctgtttcc 14220 ttatagaaac tgtggggaga atgagattca caaaggcaca tcaacaaagg tagcttgtcc 14280 tctaaggtaa atgctgactt taaaaaaatg tctcagatct atcgtttttt tacttttttc 14340 ttttt tacaa acatttattt atttggctgt gccgcatctt agttggggca ttagggatct 14400 ataatctttc agttgcagca cgtgggatcc agttccctga ccagggatcg aacctggatt 14460 ccctgcattg caagcaccga gtcttagcca ctagaccacc agagaagccc ctacttttcc 14520 ttttgaaaaa ttccaatctc cagaaaagtt gaaaaaaata gtacaataaa cacctttaac 14580 cttattcatc aatgttagca ttttaccact ttgcttttct caatgctcta cacatacatg 14640 catatatata taatatgtat tcattttttc ttgctaaatc acttggaagt aggctgaaga 14700 tattctgaca cttcactcct aaatatttta gcatacatct ctgaagaaga aagatactct 14760 cctacatgat caaaatataa tatcacagtt aagaaaatta acaccaactc attatcaccc 14820 agtaaatcca ttcagggtta atgtactgtc cttggtttca catctcttta gtttcttttc 14880 atctggaaca gtctccccac cttttttgct tttcttttat tgaatttttt gaagagtagt 14940 tcagtgttct ttgtagaatg ttccatattt tgattgtttc ctctttatta gattcaggtc 15000 aaacattctg gcaagaatac cacatagtat tacaacatgt aatgttaaat tgttctagta 15060 atggtgatga taaattagat cacttgctta aagtagtaac agctagctat ctccattact 15120 tagaaaattt tccctgtgca atctgggctt cccaagtggc tcagtggtaa agaatctacc 151 80 tgccaatgta gcagactggg gttcaatcct tgggttggga agatccctta gagaaggaaa 15240 tggcaacaca ctccagtatt cttgcctggg aaatcccaca gacagaggag cctggtgggc 15300 aatagtccat ggggtcacaa aagagtcagg catgattgag caactaaata acaacaaaat 15360 cagtaagtat tctgtgggat aatactttga cattatgtga atattctatt caacaacaac 15420 ctattaccca atggttttag catctactga taatcctttc ctgaatcaat gacattcctc 15480 agggtgaagc tcttaagtga aatttttatt tatgtattgg atgcaaaata ttttttgaga 15540 atatgtttct gaaagtatca cagaaatcaa tggaaaaagc tttgatctat aactatgttt 15600 ttgctctaca aagtcttata aagccttttt atactaacca catgtttata atgcaggttt 15660 catgaacaaa gagtttcatg gtcaaagaag ctttgaaacg ctatctaggt cctgcctgca 15720 agtggctacc aacctttaag ttccacaaga ggaatcatat cagttttact taccctggca 15780 ccggacacat agatcttcaa taaatatttg ctgaatcaaa gaataaaagg ttctgtaaga 15840 tattacagag aaaccagttt tttttcccaa aagaaattaa agtaagaaat taaggagaga 15900 gctatgctaa taatcactac ctaagaggtg tggcaggaaa aaacaagacc ctgcttgtag 15960 ccttagaaaa caattacgag agaggctgga aacaacttgc tgtgaccaaa caccac aagg 16020 agggcataat taagcattag tttaggatca gacctttgaa agcatctgtt gctccaggag 16080 cacagttctt gtcaggatga aatttcaggg caagttttcg gtaagctttc ttaagctctt 16140 catcactagc atctcgtgaa actcccagaa tttcatagta atttctgcat ttcttgatcc 16200 taaaaacaca aatcagaaac atttttttag tatgtctaca aaggtccctt ttgtacaatt 16260 aactgctaaa tagtaaatca atatccaatc tgtgagaccc aaagcagaaa gagagaaaac 16320 tggtgaacac attaatgaat cattcaattc actaacaatg tcctcctccc agtgacagct 16380 taattccttt caaagtattt ttacatctaa tatgttattc tttctatatc cctgtaagtg 16440 ggtagaacaa gtatactaat ccttatatta aggcccaatc taaggtcctc aaagttgaac 16500 cacaggacag caaaggcaga agaaataccc taaccatttt ttaaaatgta cctctttttt 16560 atggctatct ttttaaactt tgttttaaaa agttataaag gcaccttacc tacctcctga 16620 gaaatctgta tgcaggtcaa gaaggaacag ttagaaccag atatggaaca acagactggt 16680 tccaaattgg gaaaagagta tgtcaaggtt gtatattgtc accctgttta tttaacttat 16740 atgcagagtg catcatgcaa aatgccaggc tggatgaagc acaagctgga atcaagtttg 16800 cccggagaaa tatcaataac ccttagggca gaaagcgaag aagaactaa a gagcctattg 16860 atgaaagtga aagaggagag ggaaaaagtt ggcttaaaac tcaacattca gaaaactaag 16920 atcatgggat ccggtcccat cacttcatgg caaatagatg gggaaacaat ggaaacagtg 16980 agacttggcactt c ataattc ggaaacagtg 16980 agacttggcact t tt aagatgtttg ctccttggaa gaaaagctat gaccaaccta gacagcatat taaaaagcag 17100 aaacaacaaa attctgtaaa gcaactatcc ttcaataaaa aattttttta aaaagcagag 17160 acattacttt gccaacaaag gtctgtctag tcaaagctat ggtttttcaa agagtcatat 17220 atggatgtga gagttggact ataaagaaag ctgagtgccg aagaattgat gcttttgaac 17280 tgtggtgttg gagaagactc ttgagagtcc cttggactgc aaggagatca aaccagtcaa 17340 tcctaaagga aatcagttct gaatattcat tggaaggact gatgctgaag ctgaaactcc 17400 aataatttgg ccacctgatg tgaagaacag actcattgga aaagacccag atgctgggaa 17460 agattgaagg cataaggaga aggggaagac agaggatgag atggttggat ggcatcactg 17520 actcaatgga catgagtttg agcaagctct gggagttggt gatagacagg gaagcctggc 17580 atgctgcagt ccatggggtt gcaaagagtt ggacatgact gagcaactga attgaactga 17640 tggtagaaga aaaatatcca aaatactaaa ctatatacag tgaaacaact tcctgcttcc 17700 ccattttctg ttccccaatc tcctatgcta agaataacca cagttagaag ttttctgtgg 17760 ggatttcctt ggtggcctag tagttaggat tctgagcttt cactgccata acccaggttc 17820 aatccctggc cagggaactg agatcctaca agatgcacag ccaaaaaaaa aaaaaaaaa a 17880 aagttttctc attcccatgt acagagacag cacatgtata aaaacagccc cattttttaa 17940 ctttttttct tattaatacc taaatgtaag catattacaa tcacccttct acatcctttt 18000 ttcaaacaac aatttatctt ggggattgac cagtaactta cagagatgag aagtacaggg 18060 ttaatagttt ataccaccag agcagcagtt ctgacctttt tgatataaaa agtcttttat 18120 actcttgagc agcagtgagg agatacccct cgtccaaggt aaggagcatt ggctgcactt 18180 cgctggagca gccgtgaaga gataccccac gcccaaggta agagaaaccc aagtaagacg 18240 gtaggtgttg caagagggca tcagagggca gacacactga aaccatactc aaagaaaact 18300 agtcaatcta atcacactag gaccacagcc ttgtctaact caatgaaact aagccatgcc 18360 ctgtggggcc acccaagaca ggcaggtcat ggtggagaga tctgacagaa tgtggtccac 18420 tggagaaggg aatggcaaac cacttcagta ttcttgcctt gagaacccca tgaacagtat 18480 gaaaaggcaa aatgatagga tactgaaaga ggacactccc caggtcagta gatgcccgat 18540 atgctactgg agatcagtgg agaaataact ccagaaagaa tgaagggatg gagccaaagc 18600 aaaaagaata cccagctgtg gatgtgactg gtgatagaag caaggtccga tgctgtaaag 18660 agcaatattg cataggaacc tggaatgtca ggtccatgaa tcaaggcaaa t tggaagtgg 18720 tcaaacaaga gatggcaaga gtgaatgttg acattctagg aatcagcgaa ctgaaatgga 18780 ctggaatggg tgaatttaac tcagatgacc attatatcta ctactgtggg caggaatccc 18840 tcagaagaag tagccgtcat ggtcaacaaa agagtccgaa atgcagtact tggatgcaat 18900 ctcaaaaacg acagaatgat ctctgttcat ttccaaggca aaccgttcaa tatcacagta 18960 atctaagtct atgccccaac cagtaacgct gaagaagctg aagttgaatg gttctatgaa 19020 gacctacaag accttttaga actaacaccc aaaaaagatg tccttttcat tataggggac 19080 tggaatgcaa aagtaggaag taaagaaaca cctggagtaa caggcaaatt tggccttgga 19140 atacggaatg aagcagggca aagactaata gagttttgcc aagaaaatgc actggtcata 19200 acaaacaccc tcttccaaca acacaagaga agactctata catggacatc accagatggt 19260 caacaccgaa atcagattga ttatattctt tgcagccaaa gatggagaag ctctatacag 19320 ccagcaaaaa caagaccagg agctgactgt ggctcagacc ataaactcct tattgccaaa 19380 ttcagactta aattgaaaaa agtagggaaa accactagac catttaggta tgacctaaat 19440 caaatccctt atgattatac agtggaagtg agaaatagat ttaagggcct agatctgata 19500 gatagagtgc ctgatgagct atggaatgag gttcgtgaca ttgt acagga gacagggatc 19560 aagaccatcc ccatggaaaa gaaatacaaa aaagcaaaat ggctgtctgg ggaggcctta 19620 caaatagctg tgaaaagaag agaagtgaaa agcaaaggag aaaaggaaag atataaacat 19680 ctgaatgcag agttccaaag aatagcaaga agagataaga aagccttctt cagcgatcaa 19740 tgcaaagaaa tagaggaaaa caacagaatg ggaaagacta gagatctctt caaaaaatta 19800 gagataccaa aggaacattt catgcaaaga tgggctcgat aaaggacaga aatggtatgg 19860 ccctaacaga agcagaagat attaagaaga gatggcaaga atacacagat gtgtataaaa 19920 aagatcttca cgacccagat aatcacgatg gtatgatcac tgacctagag ccagacatcc 19980 tggaatgtga agtccagtgg gccttagaaa gcatcactac aaacaaagct agtggaggtg 20040 atagaattcc agttgagcta ttccaaatcc tgaaagatga tgctgtgaaa atgctgcact 20100 caacatgcca gcaaatttgg aaaactcagc agtggccaca ggactggaaa aggtcagttt 20160 tcatttcaat cccaaagaaa ggcaatgcca aagaatgctc aaactaccgc acaattgcac 20220 tcatctcaca cgctagtaaa gtaatgctca aaattctcca agccaggctt cagcaatatg 20280 tgaaccgtga acttcctgat gttcaagctg ggttagaaaa ggcagaggaa ccagagatta 20340 aattgccaac atccgctaga tcatggaaaa agcaaga gag ttccagaaaa acatctattt 20400 ctgctttatt gactatgcca aagcctttga ctgtgtggat cacaataaac tgtggaaaat 20460 tctgaaagag atgggaatac cagaccacct gatctgcctc ttgagaaatt tgtatgcagg 20520 tcaggaagca acagttagaa ctggacatgg aacaacagac tggttccaaa taggaaaagg 20580 agtatgtcaa ggctgtatat tgtcaccctg tttatttaac ttatatgcag aatacatcat 20640 gagaaacgct ggactggaac aaacacaagc tggaatcaag attgctggga gaaatatcaa 20700 taacctcaga tatgcagatg acaccactct tatggcagaa ataaagagga actaaaaagc 20760 ctcttgatga aagtgaaagt ggagagtgaa aaagttggct taaagctcaa cattcagaaa 20820 acaaagatca tggcatccgg tcccatcact tcatgggaaa tagatgggta aacagtggaa 20880 acagtgtcag actttatttt tctgggctcc aaaatcactg cagagggtga ctgcagccat 20940 gaaattaaaa gacgcttact ccttggaagg aaagttatga ccaacctaga cagcatattc 21000 aaaagcagag acgttacttt gccaacaaag gttcatctag tcaaggctat ggttttcctg 21060 tggtcatgta tgtatgtgag agttggacta tgaagaaggc tgagcgctga agaattgatg 21120 cttttgaact gtggtgttgg agaagactct tgagagtcct ttggactgca aggagatcca 21180 accagtccat tctaaaggaa atcagccctg ggatttcttt ggaaggaatg atgctaaagc 21240 tgaaactcca gtactttggc tacctcatgt gaagagttga ctcattggaa aagactctga 21300 tgctgggagg gattgggggc aagaggagaa ggggacgaca gaagatgaga tggctggatg 21360 gcatctctga ctcgatggac gtgagtctga gtgaactccg ggagttggtg atggacaggg 21420 aggcctggcg tgctgcgatt catggggtcg caaagagtcg gacacgactg agcgactgat 21480 ctgatctgat ctgatctgaa aaagatataa ttggatatta tatagcctct cacgtgaaaa 21540 aggcaaatat gtgtttgtat tattatgaag atattgtgtg tttttttggg gggtggggga 21600 cagctgcacc aagcagcatg tggaactttg ctgatcagtg gtcaaaccta caccctctgc 21660 agtggaagca aagaatctta accactggac cactagggaa gttcacaagg atagtttaaa 21720 acttgtagac ctctttccaa aatggtcttg ggattctcaa accacacctt tgagaatgac 21780 tacagcatga actctggagt caaaatgcct tggttggcat ctgactttgc tgtggactta 21840 actctgtgaa ttgttttatc ttttgaaatg ggctaataat tatacctgta tttcacagag 21900 ctgttataat taaattaagt gttcattaaa atgcttgata gcatctgaca tgatgtaaat 21960 attggctatt gttaaaaatc taccacattc tacagatggc caacaggcac acaaaaagat 22020 ctcaacattg ctaattactg ga gaaatgtg aatcaaaact acaatgaggt atatatcacc 22080 tcacattggt cagaatggcg atgatcaaaa agtctacaaa caaatgcttg agtgggtatg 22140 gaaaaaagga aaccctccta cattcttggg aggaatataa attggtgcag ctatatggag 22200 aacagtatgg tggttcctta aaaaagctaa aaatagaaca tatggtcctg gacatacatc 22260 tggaaaagat gaaaactcta atttgaaaac atacatgcac tccagtgttc acagcagcac 22320 tctttacaat agccagatga atggatagaa aagatgtggt gtatatatat atatatatat 22380 atatagtgga atattactca gccataaaaa aacaaagaaa tgccatttgt agcaacaagg 22440 acggatctag agattatcat actaggtgag gtaagtcaga caaagccaaa tatcatataa 22500 tatcacttgt atgtggacta taaaaagatg acacaaatga gctgatttac aaaacagata 22560 cactcactga cacagaaaac aaatttatgg ttaccagagg ggaaagctgt ctaggttggc 22620 cataacttcc cttccaagaa gtaagcatct tttagtttca tggctgcagt caccatctgc 22680 agtgattttg gggtccccta aaataaagtc cgccactgtt gtcactgttt ccccatctat 22740 ttgccatgaa gtgatgggac cagatgccat gatcttcgtt ttctgaatgt tgagctttaa 22800 gccaactttt tcgctctcct ctttcacttg catcaagagg ctctttagtt cttcttcact 22860 ttctgccata agggt ggtgt catctgcata tctgaggtta ttgatatttc tcccggcaat 22920 cttgattcca gcttgtgctt catccagccc agtgtttctc atgatgtact ctgcatgtaa 22980 gtgaaataag cagggtgaca atatacaacc ttgacgtact ccttttccta tttggaacca 23040 gcctgttgtt ccatgtccag ttctaactgt tgcttcctga cctgcataca gatttctcaa 23100 gaggcaggtc aggtggttgc tattcccatc tctttcagaa ttttccagtt tgttgtgatc 23160 cacacagtca aaggctttgg catagtcaat aaagcagaaa tagatgtttt tctgaaactc 23220 tcttgctttt tccatgatcc agcagatgtt ggcaatttga tctctggttc ctctgacttt 23280 tctaaaacca gcttgaacat ctggaagttc atggttcatg tattgctgaa gcctggcttg 23340 gagaattttg agcattactt tactagcgtg tgagatgagt gtaattgtgc ggtagtttga 23400 gcactctctg gcattgcctt tcttgggata ggaatgaaaa ctgacctttt ccagtcctgt 23460 ggccactgct gagttttcca aatttgctgg catattgagt gtagcacttt cacagcatca 23520 tcaaggtctg tctagtcaag gctatggttt ttccagtagt catgtatgga tgtgagagtt 23580 ggactgtaaa gaaagccgag tgctgacgaa ctgatgcttt tgaactgtgg tgttggagga 23640 gacttttgag agtctcttgg actgcaagga gatccaacca gtccatccta aaggagatca 23700 gacctgag tg ttcattggaa ggactgatat tgaagctgaa actccaataa tttggccgcc 23760 tgatgtgaag aactgactca ttggaaaaga tcctgatgct gggaaagact gaaggaggga 23820 gaaggggatg acagaggatg agatggttgg atggcatcac tgacttaatg gacatgagtt 23880 taggtaaact ccaggaattg gtgatggaca ggtaggcctg gtgtgctgca gtccatgagg 23940 tcgcaaagag tcagacacga ctgagcgact gaactgaact gaactgaaat gaggggaaag 24000 caggggtgag ggagtgggaa ggaaggggac aggttaggac ttggggatta acagatacat 24060 actactttat ataaaataat caataaggtc atactatata tatatatagc acaaggaact 24120 gtactcaata tcctataata atctataata gaaaagaatc tgaaaaagaa catatatata 24180 tatatatata tatatatata tatatatata tatatattca gttttacgta acataatatt 24240 ttgctccaca ctagaaacta acacaatttc taaatcaact atacttttaa aatctaccac 24300 attcttctaa agggtgcata ttcttgcact gttatggatg tatgggcttc ccaggtggtt 24360 cagtggtgaa gaatccacct ggcaatgcag gagacttgcg tttgatccct gggtcaggaa 24420 gatccccttg aggaggaaat ggcaccctac tccagtgttc ttgcctggaa aaaccccacg 24480 gactgagaag cctggcaggc aacaatctac agggttgcaa agagtcagac acaactgagt 24540 gactgagcat gcatgcatgt acattcccta ctgatgaggc atttaaagta cctccagttt 24600 tttctatcac acaactcagt tgagcaggca gaggctcaca gtgttgctga agagttcata 24660 agttagtaac acagtcggga ctgttcatgc agggtctgga atgtcagttg gggtccttcc 24720 ccaatagctg cttgtgggga tcctatggat tccacaccct agtttccaat tcctgattcc 24780 atggcaaata ctcccctgcc tctttctctt tccatctctc ccttatcaga tatataattt 24840 gaaaatattt tctcccattt tgtgggttga tttttcacct tcttggtggt atcttttgaa 24900 gtacaaaatt tttaattttg atgaagtcca atttgcctat attttctttt ataatgtata 24960 ctttggttgt caacacaaaa tgactcccta atccaatgtc acaaagcttt acatttatgt 25020 tttcctctaa gagttttata gttttagctc ttatatttag gtcctaggtc ttagaataaa 25080 tttttttatt tgtgtgagga aggagtccaa gttcattttt tgcaatgtgg atatccagtt 25140 gtcccaacaa gaagactagt ctttccccca ttaaattgtc ttggcactct tgtcaaaaac 25200 caattaatca taaatataag ggtttatctc aggagtgtca attctacttc attgatctaa 25260 atgcctatcc tatgctagtg cacactatct tcattctgaa tctttgcagt aggttttgaa 25320 attgggaagt atgagtcttc caactttgca cttttttccc aagactgttt tggcttccc t 25380 ggtggctcag atagtaaaga atctgcttgc aatgtaggag acacaggttc aatccctgag 25440 ttgggaagat cccctggaga agggaatggc tacccactcc agtattcttg cctggataat 25500 cccatgggca gagaagcttg gcgggctaca gtccatgggg gtcacaaaga gttggacatg 25560 actgagtgac taacacttgg ttactctagg tcctttgcat tttcaggatg catcctgaaa 25620 tccatgaatt tcaggatcag attgtcaatt tctgcaaaaa aaaggtaact gggaatttaa 25680 tagggattgt gttgaatctg tgggtcaatt tgaggaatat tatcaaaatg aagattgctg 25740 ggaaaaatat caataacctc agatacgcag atgacaccat ccttatggca gaaagtgaaa 25800 aactaaagag cctcttgacg aaagtgaaag aggagagtga aaaagtggct taaaactcaa 25860 cattcagaaa actaagatca tggcatccag tcccatcact tcatggcaaa tagatgggga 25920 aacaatggaa acagagagag atttatcttt ttgggctcca aaatcactgc agatggtgac 25980 tgcagccatg aaattataag atgcttgctc cttagaagaa aagctatgac caacctagac 26040 agcatattaa aaagcagaga cattacttta ccaacaaagg tccatatagt caaagctatg 26100 ttttttccat aagtcatgta tggatgtgag agttggacca taaagaaagc caaccgccaa 26160 agaattgatg cttttgaact gtggtgttgg agaagactct cgagagtcac t tggactgca 26220 aggagatcaa accagtcaat cccaaaggaa accagtcctg aatattcact ggaaggacta 26280 atgctgaaac tgaaactcca atactttggc cacctgatgt gaagaccctg atgctgggaa 26340 aggtcaaagg caggaggaaa aggggaagac agaggatgag atggttgaat ggcatcactg 26400 actctacgga catgagtttg agcaagctcc aggagctggt gatggatagg gaagcctggc 26460 atgcttcagt ccatggggtc gtagagttga acatgactga gcgactgaac ttaaccaata 26520 ttaagccttt cgattcataa acacaggcaa acttcccatt tagttagagt ttctttattt 26580 cttttaatga agtttgtagt tttcaatgta caatcttaca tttgttaaat ttattcctaa 26640 gcattttatt ctttttgatg ctgttataaa tgaaactgtt ttcattttca gattgttcat 26700 tgttgctgct gctgctgctg ctaagtcttg tccaactctg cgaccccata gacggcagcc 26760 caccaggttc ccccgtccct gggattttcc aggcaagaac actggagtgg gttgccattc 26820 ccttctccaa tgcatgaaag tgaaaagtga aagtgaagcc actcagtcgt gtccgactcc 26880 tagcgacccc atggactgca gccttccagg ctcctctgtc catgggattt gccaggcaag 26940 agtactggag tggggtgcca ttgctttctc cgtgttcatt gctaatgtat agaaaaataa 27000 tgatttccct atgcaagttt tcccttacag gtgccccagg gcat agaccc attgttgcta 27060 ctggtttttc ctaagttata taccaaagac tccagaaggc ttttccttga agagtcaacc 27120 aggtggtgcc caggaccctg gggcagcagc tggctcctaa aaaagtccat gactgtgggc 27180 acctctgtct ctaccctcca tgttcgtgac gccacctggc cttctcttgc atgttggaca 27240 aaggatgaac cacaagagtt tcttatcatc actttcaaat gaggattttt ctcaaaatag 27300 cagcagcagc cacgggaagg gagggaggtt ttattcattt ccacagttta tactgaacct 27360 tttggaatta ctgtttttcc caaatttttc tctggcggtg gatgtgtcat tgagccagcc 27420 atcccctgaa gccttagtac cccaagctgc ccgccaggag ggggtcacta ccctcccgga 27480 gcttctcttg gatctctttt aaaagttccc caacaaatag gattcttgct ggaaaaggaa 27540 gcttggttaa gaaatgtcaa atatggcttt ataaataaag tcctgccctg tgatgaagtg 27600 gccaccctag cccagagaga tgctcagaat tggatatgga tgagagcgac ctgcctgtgc 27660 tgaccccagt gctctgtccc catcatcccg accctctggt ttcagtagat gtccctagat 27720 gggctgagcc atgtgcaata agaacatgga tctgctatat ttcttgacta gaattctgaa 27780 attgtatgtc tataaatata tgtttattaa gtgaaaaaaa gaaaaataac gattttcgta 27840 tattgatctt tgctgaagtc atttattagt tctaata gtt tgtgtgtgaa tattccttaa 27900 aattttctat actagatcat aacatatgtg aatacagttt tactcctttt ttccaatctg 27960 gttggcttat ttttcttgcc taattgccct agcttgaaac tccagcacaa tgttgaatag 28020 taatagctag agtgaatgta gttgtcttgt tcctgttctg tttctgatct tggagagcat 28080 caaattattc atcattaagt atgatgttag ctgtggattt tttgtagatg acctttatct 28140 ggttgaggaa gttcctttct attcctagtg tactgaatat tgttatcatg agtgttggat 28200 tttgtcaaat gctttttcta cacctattga aatgatcata tgacttttgt cttttattct 28260 attaatatgg tatattacat tattttattt tcatgtgtta aacaaccttg tactcctagg 28320 ataaatctca cttggtcaag atatataatc ctttttacat gttttaatgt ttcagaaatc 28380 tgtatccatc ttgtgatgct ttcatctgct tttggttata gggtaatact ggtctcatag 28440 aatgacttgg aaagtgttcc cttctttggt attttctata ataattaatg aaagattagt 28500 taattcttct tcagacatct ggtagaattt accagttaag tcatcggtgc ctgggctttt 28560 atttgtggga aattgaattt atccatttaa gttctctaat ttgtttgttg gcatataatt 28620 gttcacagta ttctcttaca taatcattat ttttttaaga ttggtagtga tatcccctat 28680 ttcacttctg attttagtta tctgagtctt ctcccttttt tggcttggtc agtctagcta 28740 aaagtttgtc agttttgttg ataaccaata acttttgctt ttgttgatct ctattgtttt 28800 gctatcctct atttcattta ttttcactct atattatttc cttctgtttg tttgaattta 28860 gtttgttctt ctttttctag attaggttat tgatttgaga ttttagaaaa ataaacattt 28920 atagctataa atttccttta agtattggtt tagctgtatc ctatatgttt tggtgtgttg 28980 tgttttcatt ttcaatcatc tcaaagtatc ttctaatttt ccttgtgact tcttctgtga 29040 cctatttgtt atttaggaat atgttgttta atttttacac atttgtgaat tacccaaatt 29100 tccttctgtt gtcaatttca aagaacatac actgttttat tttaatcctt ttaaatttat 29160 taaggcttgt tttatagcct aacatatggt ctactctgga taatattcca tgtatacttg 29220 acaagattgt acattctgct gttgttgagt aggatgttgt acagatgtct gttatgtcaa 29280 cctggttttc cctgattctc ttagagtaga agcagggcag ttacttcaga ttatagccta 29340 acactccact tccacctaac tttgcctctg aggccaagat gttattgtgc ttcaatgata 29400 ataataatag cagtcatctg cgctaggcac tctacataaa atcattcatc taatcccacc 29460 acaggctcac ctttgtactc caagcagttg ctcctctgta taagtggagt tcccctcacc 29520 ctgtctggtc tgactccatt ca ttctcaga cttcttttcc ttctgtccct tcatgcagtt 29580 acagcaacca caggggtcat gactttcagg aggtctgtct tctgggtatt tgtttcttct 29640 gactgcatca atgtaagctg caaaacaaca gtacctccat aagggttcac tctgacctgc 29700 aaatcaactt tgctactggc cttagctatc tgggaggctg cctataggcc agtgaatgtt 29760 tgagggccag gttagtggta tcatactgac ttttacagat ccaaatccat acaaagcaga 29820 agaaacatct tccttgagaa aacaatacag aaaatataga agcaatcatg aaaaactttt 29880 aatgtcagaa acatgaagca ttaaaatgtc catcatccct ataggcatca gccagcaaat 29940 aatcttgatt cagggccagt aaaggcactc cagaggcatc tgcccaatac cttggattta 30000 gggcagcacc acggaagtct atttctctgt ggttcatggt caggagagaa ctgttcataa 30060 catatttttt tttgttgcaa tattttccac cactaagatt taaagtagag atgagttcaa 30120 tgtttaaatg gttacagaac taatacatac aagacatttg gctagataga tcaaggaggt 30180 gtatcagtaa gacctactcc tgaccacaaa gaggttactg tttaaaggga gagaatttaa 30240 ttaatcataa tgcctgcaga gtgtgaagag ggccatagga atatagtcaa agaaggcaga 30300 atgctcatgt cctgtctaga ggatctgggt aagacaaaaa acccttttct cctgcttaat 30360 atcttagtga ctacc cgttg ctccatgtcc agatctcttc ttatgggccc agcctgtctc 30420 tttagcttca aatctcacca caaggtcctc tgtactctac cttctaccac aacaatctcc 30480 cttcagatca gcaatacctt gttttctctg ggccttagca catgctgttc cttctgcctg 30540 caaaactttc tctaagtagg tctggctagc tccagcacat ccttcatgtc atcacatgta 30600 acgctttctt catgaggcct tctctgaaac ccaggtcgtg ttgagtgcct taacagtata 30660 cccccaccaa atctacgacc ttgctgtgta gcccaggaca ttttgggctt cttttgtaac 30720 tgactgttta cctgtcaaac tccctcagta gactgagagt tctgtgggct caggggacca 30780 tgtctgtcaa atccataatg cagaatacat aataggcatt ggttcagtcc tctttgcctg 30840 actgtaggat ggaatgggct ggcctatccg tcccagtgct ggatgcaggg gcaagacgaa 30900 acccaggtct agggtcaggc agcccagttt ggaggcgagg ataagggccg ggtaggcaag 30960 ctgcgggaga ccgcgttacc ccgcccaccc tggcgcgtgc cgggcttgat agctgcaggc 31020 cctgggcggg gctgggggcc ggaggcaaca agtcgtcaca ggtgactgtg ggtatcgagc 31080 cgggtcctca gcttgagcca acgcttctgg attccatatc ccggtcccct gtcctggaga 31140 gtctcctacc tcaccaggaa ccccactcaa gcccagcatc ttctcatagg cctctgtcct 31200 tcatctcagg cccaggatct cagccctttc aagtgccttc aggagtccca tccctgattt 31260 ctccgagacc ctgccctgtc ccaccctcca gccaggacct ggtaccttcc gtccagcgct 31320 ccccgctgcc caaagtcgct gccatatcgg ttcccagtca gcaggtcggc cgagcctccc 31380 ccgctcccga ggctaaaagg g 31401 <210> 3 < 211> 10001 <212> DNA <213 > Artificial Sequence <220> <223> DNAJC18_Upstream <400> 3 gaaggggggc aaaaaagcgc gaggcatgct ggtcattgta gtccaatcca taggaactgc 60 ggctcactgg actacaatcc ccggcctgca ttgcgccagg ggtcgcagct gcagtggtgt 120 aaaggggcgc gacccttgaa aggcctgtgg aggagtgttg ctgccagttg actgttagcg 180 gctcatagct gctacggtga caactgcggg cagagctttt cctcatttgt agcgtgggga 240 tagtggcttc cacgccgcat aacagctggg aagttcaagg agacgcgatg agattcactg 300 ggaaagacat tgcaaactat gggctagagg gtgacattgt gactattttt agcctcaggg 360 atgtcctttt aaaatatacg ctgaccccct aattctgtgt aaaagtgtgt gtttggtcca 420 ggacaagttt tggtgatgtt gaacccagct ctgggaagaa tcaaggaaat taaaaggtga 480 gaattttatg tggttagaag gcagcgtctc tggcatagtt tcattagaat gtccagtaag 540 agtaaggcgt agagaaagaa gaaaaagtca ggttttggag ctggcattag attcctatat 600 ttgcttttcc ttagtcttca tccatcccag gatgtctgaa gtctccatca gacatccatc 660 cctgaagtct ccagggtccc tctgagactt ctctcctcca ccccattcgt ttactggacc 720 catccccttt tctgaagtac caggccctag gagatgagtg aaaaggaaat gaagaattat 780 ccctgcattt tcccactcaa gtttgcatgg agcctagtga agaataaaag caattcttaa 840 gtgtccttac tgattcatcc ctgcaggtga gttgggatgg ccaaaaatca ggtctgcaga 900 agtgaaacat gatcatgtac atcctcaaac aaaaaaatca cctggaattc actgagatct 960 tagtcacaaa gaccaaataa gttatgtcaa ctatgatatc aaataacagc tacttaggta 1020 gttggggtgg agtggggaga tatacagccc tagacacaaa ctgaaagggt aagttgtaag 1080 cagttccatc ccttccctgc agtcatggcc ctgggataca tgaacatcag tgtgtacaca 1140 gaaggctggg tcctgaagaa gccacttgtg ctcttcagac aaggggtctg attagggaca 1200 tcctttaggt ggttaaagga agcttatttg gtttagggag gctagacaga ctactctaga 1260 agatgaacag ggaaaggtgg aatatgtctt gaagtccagg gacactgaac tagagtcaaa 1320 gatgtcagaa taccatgaag tcagaggtat ctggttgaga gtgggcatgc tagagcatta 1380 tatgggagaa attggattga gaagagtgct atgaattatg gggctactta ggagtagggt 1440 cacagactac taaccaaact gatcacattg accgcagcct tgtctaactc agtgaaactt 1500 tgagccatgc cgtgtagggc ctcccaagat ggacgggtca tggtggagag ttctgacaaa 1560 acgtggtcca ctggacaagg gaatggcaag ccacttcagt attcttgcct tgagaacacc 1620 atgctgctgc tgctgctaag tcgcttcagt catgtccg ac tctgtgtgac cccatagacg 1680 gcagcccacc aggctccccc gcccctggga ttctccaggc aagaacactg gagtgggttg 1740 ccatttcctt ctccaatgca tgaaagtgaa aagtgaaagt gaagtcgctc agttgtgtct 1800 gaccctcagc gaccccatgg actgcagcct accaggctct tccgtccatg ggattttcca 1860 ggcaagagta ctggagtggg gtgccattgc cttctcctga gaacaccata aatagcatga 1920 aaaggcaaaa agacaggaca ctgaaagatg aactccccat gttggtaggt gcccaatatg 1980 ctactggaga tcagtggaga aataactcca gaaagaatga agagacagag ccaaagcaaa 2040 aacaacacct agttgtggat atgactggtg atggaagtaa agtccgatgc tccaaagagc 2100 aatattgcat aggaacctgg aatgttaggt ccatgaatca aggcaaattg gaagtggtca 2160 aacaggagat ggcaagagtg aacatcaaca ttttaggaat caatgaacta aaatggaatg 2220 gatttggtga atttaactca gatgaccatt atatctacta ttgtgggcaa gaatccctta 2280 gaagaaatga agtagccatc atagtcaaca aaagagtcca aaatgcagta cttgaatgca 2340 atctcaaaaa tgacacaatc atctctgttt gtttccaagg caaaccattc aatatcacat 2400 ccaagtctat gcctcgacca gtaatgctga agaagctgaa gttgaacggt tatatgaaga 2460 cctacaacac cttctagaac taacacccaa aaaagatgtc ctt ttcatta taggggacgg 2520 gaatgcaaaa gtaggaagtc aagagatacc tggagtaaca ggaaaatttg gccttggagt 2580 acagaattaa gcatgtcaaa gactaacgga gttttgccaa gagaactcac tggtcatagc 2640 aaacaccctt ttctaacaac acaaaagaag actctaaaca tggacatcac cagatggtca 2700 gtactgaaat cagattgatt atattctttg cagccaaaga tggagaagct ctatgcagtc 2760 agcaaaaaca agactgggag ctgactgtgg ctcagatcat gaactcctta ttgccaaatt 2820 cagacttaaa ttgaagaaag aagggaaaac cactatacca ttcaggtatg acataagtta 2880 aatcccttat gattatacag tggaagtata ttatacaaat agattcaagg gattagatgt 2940 gatagagtgc ctgaagaact atagatggag atttgtgaca ttgtacaaga ggcagtgatc 3000 aagaccatcc ccaagaagaa atgcagaaag caaaatggtt gtctgaggag gtcttacaaa 3060 tagctgagga aagaatagaa actaaaggca aaggggaaaa ggaaagatat actcatttta 3120 attcagagtt ccaaagaata tcaaggagag atatgaaagc cttcctcagt gatcaatgca 3180 aagaaataga ggaaaataat agaatgggaa agactagaga tctcttcaag aaaattagag 3240 atatcaaggg aacatttcat gcaaagatgg gcacaataaa ggacagaaat ggtataggcc 3300 taacagaagc aaaagatatt aagaagaggt ggcaagaata cacagaaaa a ctatacaaaa 3360 aagatcttca ttacccagat aaccacaatg gtatgatccc tcatttagag ccagacatcc 3420 tggaatgtga agtcaagtgg gccttaggaa gcatcactat gaacaaagct agtggaggtg 3480 atggaattcc agctcagcta tttcaaatcc caaaagatga tgctgtgaaa gtgctacact 3540 caatttgcca gcaaatttgg aaaactcagc agtggccaca ggactggaaa aggtcagttt 3600 tcattccaat cccaaagaaa ggcaatgcca aagaatgctc aaactagcac gcaattgcac 3660 tcatctcaca tgccagtacg gagaaggcaa tggcacccca ctccagtact cttgcctgga 3720 aaatcccatg gacggaggag cctggtgggc tgcagtccat ggggtcacta agagttggac 3780 acgactgaat gacttcactt tcacttttca cttttatgca ttggagaagg gaatggcaac 3840 ccactccagt gtcttgcctg gagaatccca gggacggggg agcctggtgg gctgccgtct 3900 atggggtcgc acagagtcgg acacaactga agtgacttag cagcagcagc agcagcagca 3960 catgctagta aagtaatgct caaaattctc caagccaggt ttcaacagta cgtgaaccat 4020 aaaattccag atgttcaagc tggttttaga aaaggcagag gagccagaga tcaaattgcc 4080 aacatccatt ggatcattga aaaagcaaga gagttccaga aaaacatcta tttctgcttt 4140 attgactatg ccaaaccctt tgaccgtgtg gatcacaaca aactgtggaa aatt cttcaa 4200 gagatgggag tgccagacca cctgacctgc ctgttgagaa atctgtatac aggtcaggaa 4260 gcaacagtta gaactggaca tgaaacaaca gactggttcc aaataggaaa aggagtacgt 4320 caaggctgta tattgtcacc ttgcttattt cacttacatg cagagtacat cacgtgaaat 4380 gccaggctgg atgaaacaca agctagaatc aagatagctg gggggaatat cattaacctc 4440 agatacacag atgacaccac ccttatggca gaaaataaag aagaagcaaa gagccttttg 4500 atgaaaatga aagaggagag agaaaaagtt ggcttaaaac tcaacattca gaaaactaag 4560 atcatggcat ctggtcccat cacttcatgg caaatagatg ggggaacaat ggaaacagtg 4620 agagacttta ttttggggag ctccaaaatc actgcagatg gtgactgcag ccatgaaatt 4680 aaaagatgtt tgctccttgg aagaaaagct atgactaacc tagacaacat agtaaaaagc 4740 agagacatca ctttgccaac aaaggttcat ctagtcaaag ctatggtttt tccgtaagtc 4800 atgtatggat gtgacagttg gattataaag aaatctgagt gctgaattta tgcttttgaa 4860 ctgtggtgtt ggagaagaac tcttgagagt cccttggact acaaggagat caaaccagtc 4920 aatcctaaag gaaatcagtc ctgaatattc attggaaagg ctgatgttga agctgaagct 4980 ccaatacttg ggccacctaa tgtgaagaac tgactcattg gaaaagaccc agatgttggg 5040 aaagactgaa ggcataagga gaaggggaag acagaggatg agatggttgg atggcatcac 5100 tgattcgatg gacatgagtt tcagtaagct ccaggagttg gtgatggata gggagacctg 5160 gtgtgctgca gtccatgggg tcacaaagag ttggacatga ctgagcgact gagctgaact 5220 gaactgaact aggagtaggg tccaggttca agaattctgg acaagatttg ggaagctgag 5280 gcctcttgga aatgtctggt acctcaatga cctttcccgt tatcctctga aacctcactt 5340 cctccacttc tagatggctt ggtgacttct agatcagtaa aaaatggttt agcatacaac 5400 tttctcagat attaccatgc aaatatatcc atcagctcat ttacacttca attaacactt 5460 ctgggtgaat ttttctgtga ctatatcacc aggactaact tagaactcca aatcctcaac 5520 atttagaaca cgtctgtgcc agtttttgct taagcacgcc aactacacaa caaaaataat 5580 tcgcactgtc aaattcaatc tctgtttggt aaaagaggaa actgtgggct taaagggctc 5640 aggatggagg tgctggggaa agtaatcagc aggaagagtg agagcacctg ggaaggaaat 5700 aagggacaag aacttggcct caaacttgaa cctcaaagaa agggaggtag gtgcagaaag 5760 aagttcaaag agatgagaaa ggacatccac aaatcaaggg ggaaaatgct gagcatagcc 5820 acacaatgtt tcctagcaaa ttcaatagaa ttgagtagta ccagcctgcc ttgcctggaa 5880 aatcccatgg gcggaggagc ctggtaggct gtagtccatg gggttgctga gggtcaggca 5940 cgactgaatg acttcacttt cacttttcac tttcctgcat tggagaagga aatggtaacc 6000 cactccagtg ttcttgcctg gagaatccca ggggcggggg agcctggtgg gctgccatct 6060 atggggttgc acagagtagg acatgactga agcaacttag cagcagcagc agcagcagcc 6120 tggctttggg aaattgtgtt caagacctgg gctgggaggt ctgcagtaac tattcatctg 6180 ctaaaccact tcacctagaa tcagcctcag attttctatc agtttctcta cttttagaaa 6240 aatttgttga actgtagtga atttttgctg gtgatactga ggtttgctgt gtcccctata 6300 tgattgagat gctgctccat ccagccctgg aagaaagata ctgaataaat catccttctg 6360 tttgatggag cctgtgctcc tgagatgact tttcagttga cctcccattg gatgcaggga 6420 gaagcaaggg aagaacacat agcaagtagc ccacccctgc cacctgttcc tgccttggag 6480 tcctggagtc ctatgttagc acagggagga agccgtccag taaatgacca ccatcctgtt 6540 gtctagacat caaggaagaa atatgggctc agggaagctg cagggaaagg ctatacttgg 6600 atttcctgtc agaggtgggg ttctgtattt acataaatag taggattctc cagcagaaac 6660 ccctctaata tagcttgcta aagagcagaa ttcagcatga agtgaattca tccttaacct 6720 cttcca aaag ggtatttatt tgacaacagg aaagtcaact tggctaatgt ggtatttcaa 6780 tgtcaatttc agggtaaagg cttttagtct cttgttgata ccagttcacc ccatctctcc 6840 cacatataaa ttccttagga ctaacagctc atatttgcat aacattgtcc agaatcaggg 6900 ggaagtttgt ggccttataa tgaagatgtc aactgagtta cctagaatga aagcatgtcc 6960 ttaacctaat tagtccagca tgataaggaa ctgaaccaag tccccctctc agtcagagta 7020 acaataacaa ttcaaaaaga atccagtact tctagacatt gttaggtcac ctggagagtt 7080 ctgtcccagt gaatggattt ttgtactgga actggctagg aagtcatgtg ttttttgcat 7140 tgtggcttaa ataaaatata tcagttattt ttaaaagcta tagaattgct gtcacatgtt 7200 gggtactcca ggaaatagag tctctgagat ttgtatgctg aaggtttatt gttgagtgtt 7260 cctaacatta agtttcgagt tataagggaa gcaagattaa gcagagggag aaattgatat 7320 aaggagtcca ggtcttcacc cctacctcca cccccattga ctggtcataa gtcagctatg 7380 agccctcagc agccaacact cccagcagtt taaggtcaag agtcttgatc ctgaagtggg 7440 acctgggtag tggtactcca cattatctgc tgcagttgcc agctagccct tttactttcc 7500 aagtgtcttc acatttacta ttccatttta cctaccaagt agcctgtgag atagacaggg 7560 ctagaccagg a tcattcttc cacattgatg cacaactaac acgtaggaaa atctaagtcc 7620 cagagtcacc caatacttgt ctctggctcc cacccagaac gatctctgct caaaatattg 7680 tgttcatttt ctcttacatc ttgagtttat caagtgtaga atataagaac taaaaaagat 7740 agggaagacc aagtagtcta cccacctcac tttaaaagta agaaaatagt ttcagagatg 7800 gggatggttt cacccaaatc aagctgcaag tctgtggcag agcccagcct tttgactaga 7860 ttctctgcac tcacaagcca gttttcgtga cctctgatgt actgggcacg gtgtaccaca 7920 tagaagctac agacatgagc aggttacagg ccctgcccta gaggatttta taatccagtc 7980 ggtttcttca gaccacttta atagtttcat tgagacaagt agagtgaaac tttctgttct 8040 aagctttctc tatcctgtta cttcatagag tctaaatcca gaagaaaata gcctatggtt 8100 gcttcagaat ggctgatttg tattttcctg tgttctaaac tgccatcaat cagatgctta 8160 cggtgctcca tgcagacata cccagatgac tcttccattg actctcccca accaagattc 8220 acttccaaac cagtgtcacg gtagagatta ctaagaatac agattacttt ctttttctca 8280 ttctctttaa atgactttac tgtgtttgtg aaccttcatt ctgtttctca gacacagaaa 8340 tcatagctgt gtggggcaga gggggtagat ttaggtttat gaagtcatag atctatcctt 8400 cccagtgcta ttcccac cgg taggatgaaa gtgaatcacg tctacttggg gcaggcagag 8460 gacatttgcc tttatattat tcagctcttg tcaccgaagt gtctgaatca aaatgcgttg 8520 tacctctcgt gttctcactc tgcagaccaa ctggtgacag ttggtcactg tttatagctc 8580 cttttctgtc ccttctattg gagtggttag tcctcatcac tacttaataa atcaattaga 8640 acaaggcaga cctaataaaa cattttctcc acttcccaag tttctcagag agctccctac 8700 aggggtagaa attgtacaca tgccttacct ccaaaaggag gccccagaac tgctaaaaca 8760 ctaagaaata ctcatgggag catagacctt aaaagaaatt agactgcagg tcagaaaata 8820 actttaatgc caaggaaaaa ggatcagttc attttgtcat tctccttgct ctggctccat 8880 agactccttc ctgtgacttt gaaatggcag ggaggagcag gctgtgtttc taggctgact 8940 gttcacatgg gaaagcgtct ctcatactgg cagctccctc tcagatctgg agggatgcta 9000 gtgtcctttc agtctgggag taaatgggaa acaataattt atatctggtt cattgcctct 9060 actgattcca tcaccttctt ccttacagtc acagggcagt tgcatcatca tcagatccat 9120 tgggacccaa ggttgctttt aaagaacctg caaaataaat acataatgag gatgggtgac 9180 gttattagct atggttgggt agggaaagaa gagtctgggc cttcgagagt atctcctttc 9240 tgattaagga gatagagcct ga aaggagga ggaaggagaa ggaaggaacc aggggaaaaa 9300 tgcaccttct ctgctgaggg gcatcctttg ctgttattca tgttgatgtt cttcatcgag 9360 ataagagtga tgctctcttt ctctccattg gctgtggaac cgctggggaa ggaagggaac 9420 atgggtttta gagagaagaa gccacagggc attgagggtc aaggaattgt tggggggagt 9480 attgaaactt agacctttgt tttgagaagc ctttattttg gtggtagact gtcatcaaat 9540 gagtatgtgc tttgaagccc aaaagacttg tgtttaaatc ttagttttaa atcttactac 9600 ttgtgtgacc tggaccaagt taaaacctcc cctgcatact aagttgcttc agtcgtatct 9660 gattctttgt gaccctgtgg agtgtagccc actaggctcc tctgtccatg ggattctcca 9720 ggcaaaaatg ctggagtggg ttgccatttc ctcctctagg ggattttccc gactcaggga 9780 tcaaacacat gtctcatgtc tcctgcattg gtaggcaggt tcttcacctc tagcaccaca 9840 acttctctta agtcttattt ttttaatgta gatatttctg cattaaaatg tagaatgcag 9900 aatgtgtaaa atgcagaaat atctgccttg aagaactcta aggacttgac ataatattca 9960gttcagttca gtcgctcagt tgtgtccgac tctttgcaac c 10001

Claims (3)

A single nucleotide polymorphism (SNP) site of the DNAJC18 gene downstream, single nucleotide polymorphism sites at positions 922, 6511 and 7378 of SEQ ID NO: 1;
Single nucleotide polymorphism (SNP) sites of the DNAJC18 gene, single nucleotide polymorphism sites at positions 12439, 14758, 16701, 25415, 28427, 29222, 30021 and 30051 of SEQ ID NO: 2; and
As a single nucleotide polymorphism (SNP) site of the DNAJC18 gene upstream, 1193, 1301, 2072, 2073, 2588, 2971, 4246, 4319, 4507, 4521, 4626, 4794, 4902, 6367, single nucleotide polymorphism sites at positions 7164, 7603, 7971, 8149 and 8301;
A kit for selection of heat-resistant AHF (African Humped Cattle) breeds comprising a primer or probe that specifically binds to 10-100 consecutive nucleotide sequences including at least one single nucleotide polymorphism site. In a biological sample of cattle to provide information necessary for the selection of heat-resistant cattle AHF (African Humped cattle) breeds,
A single nucleotide polymorphism (SNP) site of the DNAJC18 gene downstream, single nucleotide polymorphism sites at positions 922, 6511 and 7378 of SEQ ID NO: 1;
Single nucleotide polymorphism (SNP) sites of the DNAJC18 gene, single nucleotide polymorphism sites at positions 12439, 14758, 16701, 25415, 28427, 29222, 30021 and 30051 of SEQ ID NO: 2; and
As a single nucleotide polymorphism (SNP) site of the DNAJC18 gene upstream, 1193, 1301, 2072, 2073, 2588, 2971, 4246, 4319, 4507, 4521, 4626, 4794, 4902, 6367, single nucleotide polymorphism sites at positions 7164, 7603, 7971, 8149 and 8301;
A method of detecting a heat-resistant cattle AHF (African Humped cattle) breed-specific marker, comprising the step of detecting one or more single nucleotide polymorphisms in one. The method of claim 2, wherein the biological sample is whole blood, plasma or serum.

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