The demand for novel strains has been rising in the domestic market to increase the production of sclerotia from Wolfiporia hoelen. To improve strain breeding efficiency, we investigated whether single-nucleotide polymorphisms (SNPs) in the RNA polymerase II subunit (RPB2) gene, which may be linked to the mating type locus, are useful for distinguishing monokaryons from dikaryons in Korean W. hoelen strains. We designed a specific primer set to efficiently amplify a region of RPB2 using PCR with the genomic DNA of 12 cultivated strains and 31 wild strains of W. hoelen collected from Korea. Nucleotide sequences of the PCR-amplified RPB2 genes were determined and analyzed for the presence of SNPs among the 43 W. hoelen strains. Previously reported SNP loci were detected in the RPB2 gene of all W. hoelen strains tested. However, these previously reported SNP loci could not be applied to differentiate monokaryons from dikaryons in approximately one-third of Korean wild strains with homozygous genotypes. Three additional SNPs in the RPB2 gene, which may improve the ability to distinguish monokaryons from dikaryons, were identified by searching through the multiple sequence alignments of the 43 W. hoelen strains. The applicability of these three novel SNPs, together with the previously known SNPs, in the RPB2 gene to W. hoelen strain breeding was verified by examining the hybrid strains and their parental strains.

밀은 세계 3대 주요 작물이지만 우리나라는 대부분 수입에 의존하고 있으며, 자급률이 1%도 도달하지 못한 실정이다. 국내 밀 품종은 40여 종 이상으로 지속적으로 개발되고 있으며, 품종마다 용도와 농업적 특성이 달라 목표 형질에 맞는 적절한 품종을 선택하여 재배하는 것이 중요하다. 품종을 정확하고 빠르게 판별하기 위해선 분자 마커 기술이 필요하다. 분자 마커는 생육 환경과 시기에 영향을 받지 않고 다양한 품종을 신속하고 정확하게 구분할 수 있어 유전적 변이성, 농업 형질 등을 분석하기에 유용하다. 본 연구는 기존에 보고된 국내 밀 32품종에 대한 SCAR (Sequence Characterized Amplified Region) 판별 마커를 재검정하여 재현성이 높은 마커를 선별하였으며, 기존에 검정되지 않은 국내 밀 9품종에 추가 적용하여 비교분석하였다. 15개의 마커 세트 중 6개의 마커가 재현성과 정확도가 높은 것으로 확인되었고, 최종적으로 국내 41품종 중 4, 7, 8, 10, 11, 12번 마커를 이용하여 다홍, 금강, 밀성, 조은, 수강, 한백, 조광, 영광을 판별할 수 있었다. 또한, 4번 마커를 통한 증폭산물의 염기서열 분석을 통해 SNP (Single Nucleotide Polymorphism)를 발굴하여 ‘올밀’에 대한 새로운 품종 판별 마커인 SdHRM1과 SdHRM2를 개발하여 HRM (High Resolution Melts) 분석을 시행하여 육안으로 판단하기 어려운 SNP를 정확하게 판별할 수 있었다. 품종간의 SNP를 활용한 품종 마커 기술은 다양한 농업형질에 적용할 수 있으며, 분자 육종 프로그램에 실질적인 도움이 되는데 큰 기여를 할 것으로 사료된다.

This study identified single nucleotide polymorphisms (SNPs) that affect the body weight of chickens. Analysis of body weight showed that the Cornish breed had the highest body weight, and the Korean native chicken (Gray Brown) had the lowest body weight. TSH is composed of an α-subunit and a β-subunit, and the TSH-β gene encoding the β-subunit has been reported to be associated with obesity. In chickens, it is located on chromosome 26 and is reported to be associated with growth. The calcium-sensing receptor gene (CaSR) plays a role in the regulation of extracellular calcium homeostasis and is responsible for calcium absorption in the urinary tract, which affects the eggshell quality in poultry. It was shown that TSH-β was strongly correlated with weight in Cornish and Korean native (Gray Brown) chickens, particularly in those with the CC trait. However, CaSR showed no association with body weight in poultry; it was associated with calcium and the eggshell. Thus, selection for TSH-β can be used to produce individuals with more favorable traits in terms of body weight.

본 연구에 이용되는 국내 재래 흑염소는 유전적 자원을 보존하고 품종을 유지하기 위하여 소규모 집단에서 고도의 근친교배를 수행하고 있어 유전적 자질의 다양성이 저해되고 있을 것으로 우려되었다. 따라서 본 연구는 국내 재래흑염소 경상대계통 집단의 유전체 정보를 활용하여 재래흑염소 집단의 유전적 특성과 유효집단의 크기를 추정하고자 실시하였다. 국내 재래 흑염소의 유전체 정보는 Illumina Goat SNP 50k chip (illumina, inc., San Diego, CA)의 정보를 분석하여 연구에 이용하였다. 각 염색체의 인접 표지인자 와의 연관불평형 (Linkage Disequilibrium)은 0.225로 추정이 되었다. 또한, 표지인자 사이의 거리가 증가함에 따라 연관불평형의 값은 감소되는 것으로 나타났다. 국내 재래 흑염소의 유효집단크기는 최근의 세대로 오는 경우 감소하는 추세를 보였으며, 13세대에서 유효집단의 크기는 29두로 추정되었다. 재래 흑염소 경상대계통은 낮은 연관불평형 값과 유효집단 크기를 보여 유전적 자원의 다양성이 낮을 것으로 보이며, 이를 해결하기 위한 계획적인 교배와 품종 집단의 크기를 키우기 위한 대책이 필요할 것으로 사료된다.

To control an external parasitic mite, a honey bee line possessing high hygienic behavior (HHB) against an external parasitic mite, Varroa destructor, has been bred in South Korea and an assessment method has been necessitated to diagnose HHB line from the low hygienic behavior (LHB) line. Thus, in this study, we developed single nucleotide polymorphism (SNP) markers from whole genome sequencing of each 20 worker bees from HHB and LHB lines of A. mellifera ligustica (Hymenoptera: Apidae). An average of 319,445,977 sequence reads was mapped to the known A. mellifera reference genome (an average of 87.46%). In 2,316,128 and 3,266,756 SNPs from each HHB and LHB line, an average of 93.6% and was located in the intergenic spacers and introns, whereas, the remaining 6.4% was located in the genic region, respectively. Among them 20 SNPs that were fixed at each line possessing within-individual homozygosity were selected and each four SNPs were used to diagnose the two honey bee lines either by typical PCR-restriction fragment length polymorphism method or allele-specific PCR. The remaining six SNPs had the size difference, enabling relatively easy differentiation between the two honey bee lines on typical agarose gel and another remaining six SNPs only has sequence difference including SNP sites. Thus, these SNP markers can be used to diagnose the honey bee line with HHB from LHB line against V. destructor.

We developed single nucleotide polymorphism (SNP) markers and are establishing diagnostic systems to distinguish disease resistance- and susceptible-strains of honey bees using the SNPs. For development of SNP markers, whole genome was sequenced each from 20 individuals of “disease resistance-strain” and “susceptible-strain” of Apis mellifera ligustica using the Illumina HiSeq 2000 sequencer. Approximately, 344 and 294 million sequence reads were mapped to the honeybee reference assembly (Amel_4.5) for each strain, respectively. Among the total 2,246,428 SNPs yielded, 33 were found to be fixed between the two strains with all homozygosity. Sixteen of them were casually amplified and sequenced from randomly selected each 10 individual of honey bees from each strain and presented strain specific SNPs. These ten SNPs were used to diagnose the two strains either by original size difference, caused by indel-accompanying SNP, typical PCR-RFLP, or AS PCR.

This study aimed to indentify single nucleotide polymorphisms (SNPs) in exon region of the glycerol-3-phosphate dehydrogenase 1 (GPD1) gene and to evaluate their associations with meat yield and quality traits in Hanwoo (Korean cattle). Two SNP markers, g.2766C>T and g.5105A>G were identified in the exons 5 and 8, respectively. Genotyping of the two SNPs was carried out using PCR-RFLP analysis in 309 Hanwoo steers to evaluate their associations with meat yield and quality traits. As a result, g.2766C>T in exon 5 was significantly associated with carcass weight (CW) and marbling score (MS). Animals with the CC genotype of g.2766C>T had heavier CW than those with the CA or AA genotype. Animals with the CC genotype of g.2766C>T also had higher MS than those with the CA or AA genotype. Additive effect was also observed with CW and MS traits. We constructed haplotypes by linkage disequilibrium analysis and analyzed association between haplotypes and meat yield and quality traits. Haplotype of GPD1 gene was associated with CW. As a result, animals with CA haplotype had heavier CW than TG haplotype. These findings suggest that the SNPs of bovine GPD1 gene may be a useful molecular marker for selection of meat yield and quality traits in Hanwoo.

This study tested the association between genotypes of the single nucleotide polymorphism (SNP) marker, rs81437607 and capric acid (FA_C10_0) compositions in longissimus dorsi muscle in pigs. Eighteen fatty acid (FA) compositions were measured in a total of 974 F2 animals among 1,106 F2 progeny produced between Landrace and Jeju Black Pig (JBP). Among FA compositions tested, we identified a cluster of highly significant SNPs for capric acid compositions on 58 Mb position of Sus scrofa chromosome 12 (SSC12) using genome-wide association study (GWAS) with F2 genotypes from SNP panel analysis. GWAS results showed that the rs81437607 was the highest trait-related SNP marker with capric acid levels. Three genotypes (C/C, C/T and T/T) of rs81437607 marker were found in F2 population by further pyrosequencing. Association analysis results showed the significant differences between rs81437607 genotypes and capric acid compositions (P<0.05). The F2 pigs harboring rs81437607 C/C (0.119±0.002%) and C/T (0.116±0.002%) genotypes showed additively higher levels of capric acid content than those of T/T homozygotes (0.109±0.002%) (P=1.30×10-12). These results suggested that the genetic variations of rs81437607 may be helpful to find causative variants and assist as molecular genetic markers for improving the capric acid contents in longissimus dorsi muscle in pigs.

This study was conducted for SNPs in the 5'-regions of estrogen receptor-α(ESR-α), and association with calving interval (CI), service per conception (SPC) and 305 days milk yield in Hanwoo and Holstein dairy cattle. The genet-ic improvement was incurred low reproduction performance. The objective of this study was to investigate connec-tions between single nucleotide polymorphisms (SNP) of Estrogen receptor-α (ESR-α) with reproduction performance (calving interval, service per conception, and 305 d milk yield) in Hanwoo and Holstein dairy cattle. Hanwoo and Holstein blood samples were collected from 183 and 124 dam of breeding farms and DNA was extracted. Primer design was based on NCBI GenBank (Accession No. AY340579). The PCR-RFLP method with Bgl I was used to ge-notype the cattle. The result showed two variants of the ESR-α gene. The Bgl I cut the 492 bp amplification pro- duct into 322 bp and 170 bp fragments for allele G, while allele A remained uncut, resulting in two restriction frag-ments for homozygote G/G and three fragments for heterozygote A/G. We found two of different genotypes in the-se breeds, A/G and G/G. In Hanwoo, the A/G genotype frequency was 0.13, and G/G was 0.87. The CI of A/G was 382.18±10.03 days, and G/G was 381.69±5.22 days. The SPC of A/G was 1.62±0.16, and G/G was 1.32±0.04. While CI showed no significance difference, SPC exhibited significant difference (p<0.05). In Holstein cattle, the frequency of genotype A/G was 0.02 and G/G was 0.98. The 305 days milk yield of A/G was 7,253.00±936.00 kg and of G/G was 8,747.51±204.88 kg, showing no significant difference.

MYC (v-myelocytomatosis viral oncogene homologue) is a regulator gene that encodes for a nuclear phosphoprotein. Porcine MYC gene was mapped on chromosome SSC 4p13 and is associated with a variety of functions such as cell proliferation and cell growth. MYC expression is coupled to a multitude of physiological processes and is regulated by hormones, growth factors, cytokines, lymphokines, nutritional status, development and differentiation. MYC is also involved in myogenesis, muscle hyperplasia and adipogenesis. In this study, we investigated SNPs in MYC gene and their association with economic traits in Duroc, Landrace and Yorkshire populations. We detected a single point mutation in exon 3 of porcine MYC gene as a change of T to C at 906 base (amino acid position 302, nonsynomous mutation of alanine) in MYC-N domain. MYC mutation (T906C) was significantly associated with age at 90 kg in these breeds, signifying that this mutation can serve as a selection marker for growth traits in pigs.

3종의 초위성체 마커를 이용해 분석한 대립유전자형을 F0, F1 그리고 F2로 세대 간 구분 하여 동일한 개체 출현확률 값을 추정한 결과 F2의 무작위교배집단으로 가정한 경우 13종의 초위성체 마커는 3.84 × 10-23의 추정치를 나타내 37개의 SNP 마커를 이용할 경우와 유사한 동일개체 출현확률 추정치를 유추할 수 있었다. 본 연구에 사용한 실험축군은 2품종 상호교배로 만들어졌다. 친자감정확률 추정치를 전체집단을 대상으로 13종의 초위성체 마커와 37개의 SNP 마커를 이용하여 분석한 결과 부모를 동시에 찾을 수 있는 추정치인 PEpu의 경우 초위성체 마커는 0.97897이고 SNP 마커는 0.99149였으며, 한쪽 부모를 찾을 수 있는 추정치인 PE의 경우 초위성체 마커는 0.99916이고 SNP 마커는 0.99949로 나타났다. 또한 가능한 후보 부모들로부터 가장 확률이 높은 부모를 찾을 추정치인 PNEpp의 경우는 초위성체 마커와 SNP 마커 둘 다 1.00000으로 추정 되었다. 한정된 부모집단 내 한정된 대립유전자형을 통해 대량의 비육돈이 생산되는 국내의 양돈산업의 경우 DNA 마커의 특성, 분석집단의 크기, 유전자형 분석의 정확도와 비용, 분석된 자료 관리의 용이성 및 기존 분석 시스템과의 호환성 등을 고려하여 효율성과 경제성이 높은 마커를 선정해 마커 조합을 만드는 것이 필요할 것으로 사료된다.

Myopalladin (MYPN) is an important expression gene associated with regulation of Z-line structure in muscle and maintains sarcomeric integrity. In this study, we investigated the association between MYPN A1795G SNP (single nucleotide polymorphism) and carcass traits (LMA, longissimus muscle area; CW, carcass weight; BF, backfat thickness; MS, marbling score) in Korean cattle. The MYPN A1795G SNP was genotyped in 212 steers and analyzed the associations with carcass traits by PCR-RFLP (Restriction fragment length polymorphism) method. The allele frequencies were 0.566 for G allele and 0.434 for A allele. And the genotype frequencies of GG, GA, and AA genotype were 32.1%, 49%, and 18.9%, respectively. Association analysis indicated that the A1795G SNP of MYPN gene showed a significant association with LMA (p<0.05). The steers with GG genotype had higher LMA than those with the genotypes AA. But no significant associations were observed in other carcass traits (CW, BF, MS). The steers with the GG genotype showed higher CW and BF than those with the genotypes AA and GA. These results suggest that the A1795G SNP of the MYPN gene is associated with LMA and may be useful for candidate marker-assisted selection to increase the levels of LMA in Korean cattle.

Aldehyde dehydrogenase (ALDH) plays an important role in alcohol metabolism; ALDH is responsible for the oxidation of acetaldehyde generated during alcohol oxidation. ALDH is also known to oxidize various other endogenous and exogenous aldehydes. Cytochrome P-450 2E1 (CYP2E1), a liver microsomal enzyme, also metabolizes acetaldehyde and ethanol and can be induced by other inducers including acetone and ethanol. We examined single nucleotide polymorphisms (SNP) of ALDH and CYP2E1 genotypes in Korean. Restriction fragment length polymorphism (RFLP) method was used to determine ALDH and CYP2E1 SNP. Mutation in ALDH was 60% (heterozygote 46.7% and homozygote 13.3%) among 15 cases. CYP2E1 mutation was 52.7% (heterozygote 47.4% and homozygote 5.3%) among 19 cases.

Leaf mold disease in tomato (Solanum lycopersicum) is caused by Cladosporium fulvum, a fungal leaf pathogen. One of effective ways to control leaf mold is to breed disease-resistant tomato cultivars. Cf-4 and Cf-9 resistance (R) genes encode proteins that carry a leucine rich repeat domain and are located in plasma membrane. They trigger hypersensitive response following recognition of corresponding Avr4 and Avr9 proteins of C. fulvum, respectively. Cf-4 and Cf-9 genes are originated from wild tomato species S. habrochaites and S. pimpinellifolium and have been introgressed into commercial tomato cultivars. These two highly homologous orthologs exist as a cluster with four highly homologous paralogs. Due to this reason, development of genetic markers to distinguish these two functional R genes from their orthologs and paralogs is difficult. In this study, we tried to develop single-nucleotide polymorphism (SNP) markers to select tomato cultivars carrying resistant Cf-9 genotype. The genomic sequences of resistant Cf-4 and Cf-9 alleles, susceptible cf-9 alleles, and their paralogs were obtained from the GenBank database, and two functional SNPs causing non-synonymous substitution were found among them. Based on two SNPs, the Cf-9_2-SNP-F/R primer set for high resolution melting (HRM) analysis was developed. HRM analysis with this primer set could successfully distinguish tomato cultivars carrying resistant Cf-9 allele among 30 commercial tomato cultivars, which were characterized with the gene-based marker. These indicate that the SNP marker developed in this study is useful to trace Cf-9 genotype efficiently in marker-assisted selection in tomato.

Blackleg disease caused by Leptosphaeria maculans, is the most devastating disease of Brassica germplam worldwide that causes million tonnes of crop losses per year throughout the world. To date, a total of 12 race-specific resistance genes of Brassica napus to L. maculans have been reported but linkage mapping analysis reveals that all of those loci are located in A genome i.e., in B. rapa chromosomes. B. oleracea has high ancestral synteny with B. rapa through their evolution. We believe that presence of qualitative resistance is possible in B. oleracea germplasm. The present study was therefore planned to find out any race-specific qualitative resistance gene present in C genome of B. oleracea. A total of 16 microsatellite markers were used which are linked to seven different Rlm and Lep genes of B. napus to screen 32 inbred lines of cabbage. Primers were designed based on homology assessment in corresponding nucleotide sequence available in Bolbase (a B. oleracea genome database, http://www.ocri-genomics.org/bolbase/index.html), located in B. oleracea scaffolds/chromosomes. Out of 16 SSR markers, 13 were found polymorphic which indicates possible existence of resistant genes in cabbage lines. The inbred lines are then assessed against two L. maculans stains with known avirulent genes. Some inbred lines were hypersensitive against gene-specific virulent strains of L. maculans that confirmed existence of Rlm1, Rlm2, Rlm4, LepR3 and LepR4 in the cabbage lines. In this way we were able to select out resistant and susceptible lines against each resistant gene. The gene-specific polymorphic SSR marker regions were cloned and sequenced and candidate SNPs were identified for confirmation of their functionality.

To successful molecular breeding, identification and functional characterization of breeding related genes and development of molecular breeding techniques using DNA markers are essential. Although the development of a useful marker is difficult in the aspect of time, cost and effort, many markers are being developed to be used in molecular breeding and developed markers have been used in many fields. Single nucleotide polymorphisms (SNPs) markers were widely used for genomic research and breeding, but has hardly been validated for screening functional genes in olive flounder. We identified single nucleotide polymorphisms (SNPs) from expressed sequence tag (EST) database in olive flounder; out of a total 4,327 ESTs, 693 contigs and 514 SNPs were detected in total EST, and these substitutions include 297 transitions and 217 transversions. As a result, 144 SNP markers were developed on the basis of 514 SNP to selection of useful gene region, and then applied to each of eight wild and culture olive flounder (total 16 samples). In our experimental result, only 32 markers had detected polymorphism in sample, also identified 21 transitions and 11 transversions, whereas indel was not detected in polymorphic SNPs. Heterozygosity of wild and cultured olive flounder using the 32 SNP markers is 0.34 and 0.29, respectively. In conclusion, we identified SNP and polymorphism in olive flounder using newly designed marker, it supports that developed markers are suitable for SNP detection and diversity analysis in olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

Molecular markers are useful for selecting to include superior character genetic like as strong immune system and rapid growth in fish. The marker is also very important part of breeding technology in Olive flounder (Paralichthys olivaceus). Single nucleotide polymorphisms (SNPs) marker is already in use widely for genomic research and breeding. But this SNPs marker hardly has been validated for screening functional genes in Olive flounder. We study identify single nucleotide polymorphisms (SNPs) on Expressed sequence tag (EST) database, develop usable SNP marker and apply to wild sample and cultured of olive flounder. As a result, Out of total 4.327 ESTs, 693contigs and 514 SNP from total contigs were detected while these substitutions include 297 transitions and 217 transversions. 144 developed markers were applied in 16 samples (wild 8, culture 8), Out of total marker, only 32 markers had detected polymorphic in sample. Polymorphism of 32 markers was observed in the variety genes region involved in immunity and protein synthesis. And the 32 marker were identified 21 transitions, 11 transversions, and indel was not detected in polymorphic SNPs. The analysis on heterozygosity by sample showed 0.34 in wild sample and 0.29 in cultured sample. In conclusion, we was identified SNP and Polymorphism by designed new marker, it supports that development marker is suitable for SNP detection and diversity analysis in Olive flounder. The outcome of this study can be basic data for researches for immunity gene and characteristic with SNP.

With the development of next generation sequencing (NGS) technology, the variation of sequences represented as SNP between cultivars becomes available at genome level. The major domestic cultivars with high yield have been developed by breeding of indica and japonica, it is important to localize the region of origin according to the genotype for further characterization of unique features of cultivars. For the localization of SNP at genome level, the paired end sequences of 6 major domestic rice cultivars, Ilmi, Ilpoom, Sulgaeng, Bakjinju, Hwayoung and Woonkwang were compared against Japonica and Indica Rice Genomes as reference genomes. The genomic DNAs were prepared from callus tissues and paired-end of the fragments were sequenced with NGS Sequencer, Illumina HISeq. About 50x coverage of paired-end sequences were trimmed according to the quality of the sequences, and errors were corrected with statistical analysis of kmers of 15. The trim-corrected sequences were mapped and variants were analyzed against reference genomes. The overall change rate of Ilmi against Nipponbare IRGSP 1.0 and Indica BGI 93-11 reference genomes were 0.92 base/1kb (1/1,079 base) and 8.09 base/1kb (1 base/123 bases), respectively. Among 6 cultivars, overall rate of Bakjinju showed the lowest overall change rate of 0,53 base/1kb, and Hwayoung showed highest frequency of 0.92 base/1kb. Compared to high level in the range of change rate of 7.0-9.3 base/1kb against indica, domestic cultivars showed lower range of change rate 0.2-3.3 base/1kb with unique local high peak against japonica genome depend on the chromosomes. Compared to assembly of genome sequences, the variation of nucleotides compared to reference sequences is much faster and simple to characterize the genotype. The types of variation and the effect on functional categories will be presented.