유전자변형작물의 개발 현황 및 안전관리 동향

동춘매,김주원,강정하,박중연,김군도,공희정 한국수산해양교육학회 2017 水産海洋敎育硏究 Vol.29 No.6

Genetically modified organisms (GMOs) are created by biotechnology to improve crop production. The commercialization and commercial cultivation of GMOs are gradually increasing. The area planted with genetically modified crops, which was only 1.7 million hectares in 1996, increased by 18.51 million hectares in 2016. Compared to 1996, it has increased by more than 100 times. The country that grows the most genetically modified crops is the United States; the area of cultivation in the United States is 72.9 million hectares. The cultivation area of genetically modified crops is increasing annually in the world, in the order of Brazil (49.1 million hectares), Argentina (2.380 million hectares), Canada (1.160 million hectares), and India (10.80 million hectares). Korea has not yet grown genetically modified crops, but it has been importing genetically modified crops for food and feed since 2001. In 2016, 22.4 million tons (21%) were imported for food and 77.4 million tons (79%) were imported for feed. In addition, many genetically modified crops, including rice, pepper, soybean, and grass, have been developed by research institutes, universities, and companies, and some of these crops are expected to be put into practical use soon in Korea at the safety evaluation stage. The number of genetically modified crops has increased steadily worldwide to benefit societies, but these crops may pose risks to the ecological environment and to human health and therefore require careful regulation. This study investigated the current status of genetically modified crops, both domestically and overseas, and examined the safety management trends and labeling systems for GMOs.

Development of Species-Specific PCR Primers for the Rapid and Simultaneous Identification of the Six Species of Genus Takifugu

동춘매,박연정,노재구,노은수,안철민,강정하,박정연,김은미 한국발생생물학회 2019 발생과 생식 Vol.23 No.4

Pufferfish (Takifugu spp.) are economically important edible marine fish. Mistakes in pufferfish classification can lead to poisoning; therefore, accurate species identification is critical. In this study, we used the mtDNA cytochrome c oxidase subunit I gene (COI) to design specific primers for six Takifugu species among the 21 domestic or imported pufferfish species legally sold for consumption in Korea. We rapidly and simultaneously identified these pufferfish species using a highly efficient, multiplex polymerase chain reaction (PCR) system with the six species-specific primers. The results showed that species-specific multiplex PCR (multiplex species-specific polymerase chain reaction; MSS-PCR) either specifically amplified PCR products of a unique size or failed. MSS-PCR yielded amplification fragment lengths of 897 bp for Takifugu pardalis, 822 bp for T. porphyreus, 667 bp for T. niphobles, 454 bp for T. poecilonotus, 366 bp for T. rubripes, and 230 bp for T. xanthpterus using the species-specific primers and a control primer (ca. 1,200 bp). We visualized the results using agarose gel electrophoresis to obtain accurate contrasts of the six Takifugu species. MSS-PCR analysis is easily performed and provides identification results within 6 h. This technique is a powerful tool for the discrimination of Takifugu species and will help prevent falsified labeling, protect consumer rights, and reduce the risk of pufferfish poisoning.

차세대 염기서열 분석법을 사용한 벤자리(Parapristipoma trilineatum)의 microsatellite 마커의 개발 및 유전학적 특성 분석

동춘매,이미난,노재구,박진우,김영옥,김은미 한국생명과학회 2023 생명과학회지 Vol.33 No.8

This study was conducted to develop microsatellite markers in Parapristipoma trilineatum using next- generation sequencing. A total of 402,244,934 reads were generated on the Illumina Hiseq X Ten System, yielding 60,738,985,034 bp of sequences. The de novo assembly resulted in 1,320,995 contigs. A total of 952,326 contigs (0.016%) including 151 microsatellite loci were derived from the 1,320,995 contigs longer than 640 bp. A total of 34 primer sets were designed from the 151 microsatellite loci. As a result, 15 microsatellite loci were chosen and used for assuming population genetic parameters in the wild and farmed populations. The mean number of effective alleles was 12, ranging from 6 to 25. The observed heterozygosity (HO) and the expected heterozygosity (HE) ranged between 0.530 and 0.873, with an average of 0.750, and from 0.647 to 0.895, with an average of 0.793, respectively. According to these results, the developed set of 15 microsatellite markers is expected to be useful for the analysis of genetic characteristics in the population of P. trilineatum in Korea. There are requirements now for further genetic information, fishery resource management, breeding guidelines, support with the selection of breeds and studies on the effects of release, all of which will improve species conservation, and through future research, we aim to offer genetic foundational data with that goal.

종-특이적 PCR 분석법을 이용한 신속하고 간편한 말전복의 종판별법 개발

동춘매,이미난,강정하,박중연,남보혜,노재구,김필연,조영아,김은미 한국패류학회 2018 The Korean Journal of Malacology Vol.34 No.1

This study was performed to identify rapidly Haliotis gigantea using polymerase chain reaction with species-specific primers. Around 680 bp of the mitochondrial ND5 gene region from four Haliotis species were aligned and species-specific forward primer was designed based on the single nucleotide polymorphism (SNP) from H. gigantea. The optimal PCR conditions were selected by cross reactivity using gradient PCR method from 55°C to 66°C. Species-specific PCR (SS-PCR) amplification reactions with two pairs of primers were performed for a five specimens of Haliotis species. SS-PCR leads to a species specific amplification of a 1,006 bp fragment in H. discus hannai, H. discus discus, H. madaka and 786 bp in H. gigantea, respectively. The two different sizes of each PCR products can be quickly and easily detected by single gel electrophoresis. The sensitivity of the SS-PCR was up to 1ng/μl DNA as a starting concentration in H. gigantea tested. Therefore SS-PCR technique with species-specific primer based on SNP could be a powerful tool for discrimination of H. gigantean and can contribute to the prevention of falsified labeling of this species.

NGS를 이용한 참굴 (Crassostrea gigas) microsatellite markers 개발

동춘매,이혜민,이미난,노은수,남보혜,김영옥,김은미 한국패류학회 2022 The Korean Journal of Malacology Vol.38 No.3

This study was conducted to develop microsatellite markers in Crassostrea gigas using next-generation sequencing. A total of 46,335,655,445 bp reads were generated on an Illumina Hiseq x ten system, yielding 600,863,377 bp sequences. The de novo assembly resulted in 30.636 contigs. A total of 261 contigs, including 56 microsatellite loci, were derived from 30,636 contigs longer than 518 bp. A total of 22 polymorphic nuclear microsatellite markers were chosen to evaluate population genetic parameters in the farm. The mean number of effective alleles was 9, ranging from 3 to 25. The observed heterozygosity (HO) and expected heterozygosity (HE) ranged between 0.104 and 0.896 with an average of 0.469 and from 0.214 to 0.947 with an average of 0.579, respectively. No significant linkage disequilibrium was observed after Bonferroni revision in any loci. The results show that the 22 polymorphic nuclear microsatellite markers can be used to study the population and conservation genetics of Crassostrea gigas in Korea. The analysis of polymorphic SSR could provide an important experimental tool for examining a range of issues in Crassostrea gigas genetics

Microsatellite marker 분석을 이용한 명태(Theragra chalcogramma) 5 집단의 유전적 다양성 및 유연관계 분석

동춘매(Chun Mae Dong),강정하(Jung-Ha Kang),변순규(Soon-Gyu Byun),박기영(Kie-Young Park),박중연(Jung Youn Park),공희정(Hee Jeong Kong),안철민(Cheul Min An),김군도(Gun-Do Kim),김은미(Eun-Mi Kim) 한국생명과학회 2016 생명과학회지 Vol.26 No.11

한류성 어종인 명태는 우리나라 동해를 비롯한 일본, 러시아 북부의 오호츠크해, 베링해, 알래스카 등지에 서식하는 중요한 수산자원으로, 우리나라에서는 그 어획량이 매년 감소하고 있어, 그 자원량의 회복과 보존 및 관리가 필요한 대표적 어종이다. 그러나, 이러한 중요성에도 불구하고 국내에서 명태의 유전학적 집단 분석에 관한 연구는 많이 수행되지 않은 실정이다. 본 연구에서는 우리나라 동해, 러시아, 미국 명태 집단 및 일본 명태 집단과의 유전적 다양성과 유연관계를 분석하여 명태자원의 보존과 관리를 위한 과학적 자료를 제공하기 위해 유전적 다양성 및 계군 분석에 널리 사용되고 있는 microsatellite marker (msDNA) 8개를 사용하여 명태 집단의 유전자형을 분석하였다. 우리나라 동해, 러시아, 미국 및 일본 집단에서 채집된 총 186개체를 분석한 결과, 대립유전자수는 최소 7.13개에서 최대 10.63개로 나타났고, 평균 대립유전자의 수는 9.05개로 나타났다. 기대치와 관찰치 이형 접합율은 각각 0.698과 0.732로 조사되어, 현재 확보된 명태 집단의 유전적 다양성은 비교적 잘 유지되고 있는 것으로 나타났다. 유전학적 유연관계 분석을 위한 유전적 거리, Pairwise FST값, UPGMA와 주성분분석, AMOVA test 분석 결과, 우리나라 동해, 러시아, 미국의 명태 집단 간 유전적 차이는 거의 없었으나 일본 명태 집단과는 낮은 수치이지만 유의한 유전적 차이가 있음을 확인하였다(p<0.05). 본 연구에서 확인된 유전학적 분석을 통한 명태집단의 유전적 특성 및 주변국 집단과의 유연관계 분석결과는 우리나라 동해의 중요한 수산유전자원으로서의 명태에 대한 중요한 과학적인 근거자료가 될 것이며, 앞으로 명태 자원의 보존, 평가 및 이용에 활용 가능한 정보를 제공할 것이다. A comprehensive analysis of the genetic diversity and relationship of the cold-water fishery walleye pollock (Theragra chalcogramma), the most abundant economically important fishery resource in the East sea of Korea, has not been carried out, despite its importance in Korea. The present study assessed the genetic diversity and relationship between five walleye pollock populations (Korean population, Russian population, USA population, and Japanese populations) of T. chalcogramma using eight microsatellite DNA (msDNA) markers to provide the scientific data for the preservation and management of the Pollock fishery resource. The results of the analysis of 186 individuals of the Pollock revealed a range of 7.13–10.63 numbers of alleles (mean number of alleles=9.05). The means of observed heterozygosity (HO), expected heterozygosity (HE) were 0.732 and 0.698, respectively. The results of genetic distance, Pairwise FST, UPGMA (UPGMA: un-weighted pair-group method with an arithmetical average) (the phylogenetic tree), PCA (PCA: Principal Coordinate analysis) analysis pointed to significant differences between the Korean population, Russian population, USA population, and Japanese populations, although small (p<0.05). These results shed light on the genetic diversity and relationships of T. chalcogramma and can be utilized for research on the evaluation and conservation of Korean T. chalcogramma as genetic resources.

차세대 염기서열 분석법을 이용한 방어(Seriola quinqueradiata)의 microsatellite 마커의 개발 및 유전적 특성 분석

동춘매(Chun Mae Dong),이미난(Mi-Nan Lee),김은미(Eun-Mi Kim),박중연(Jung Youn Park),김군도(Gun-Do Kim),노재구(Jae Koo Noh) 한국생명과학회 2020 생명과학회지 Vol.30 No.3

본 연구는 차세대 염기서열 분석법(NGS)을 이용하여 방어의 microsatellite 마커를 개발하고, 개발된 마커를 이용하여 방어 집단의 유전적 특성을 분석하기 위해 수행되었다. 차세대 염기서열 분석 장비인 Illumina Hiseq2500를 이용하여 총 28,873,374개의 read들을 얻어 assembly를 수행한 결과, 전체의 약 1.6%에 해당하는 466,359개의 read들이 assembly 되었으며, 이 read들의 총 길이는 7,247,216,874 bp로 확인되었다. 크기가 518 bp 이상이 되는 contig는 30.729개로 나타났으며, 이 중 microsatellite 영역을 포함하는 contig 132개(0.43%)를 1차로 선별하고, PCR 증폭 여부 및 유전자형 분석을 통해 microsatellite 후보 60개를 2차로 선별하였다. 그 중 방어집단의 마커로서 유용한 15개의 microsatellite 마커를 선택하였다. 방어집단을 대상으로 개발된 15개의 microsatellite 마커로 분석한 결과, 관찰된 유효 대립유전자수(NA)는 평균 18.5(11~30)로 나타났다. 평균 관측치 이형접합도(HO)와 평균기대치 이형접합도(HE)는 각각 0.812(0.431~0.972)와 0.896(0.782~0.949)으로 나타났다. 다형성이 관찰된 모든 microsatellite 마커 간의 연관불평형은 나타나지 않았으며, 해산어의 평균 HE 값인 0.79 이상의 수치를 나타내었다. 따라서 본 연구에서 개발된 15개의 microsatellite 마커는 방어 집단의 유전적 다양성 분석에 유용할 것으로 사료된다. This study was conducted to develop microsatellite markers in Seriola quinqueradiata using next-generation sequencing. A total of 28,873,374 reads were generated on an Illumina Hiseq2500 system, yielding 7,247,216,874 bp sequences. The de novo assembly resulted in 466,359 contigs. A total of 132 contigs (0.43%), including 60 microsatellite loci, were derived from 30,729 contigs longer than 518 bp. A total of 60 primer sets were designed from the 132 microsatellite loci. A total of 15 polymorphic nuclear microsatellite loci were chosen to evaluate population genetic parameters in the parents and offspring. The mean number of effective alleles was 18.5, ranging from 11 to 30. The observed heterozygosity (HO) and expected heterozygosity (HE) ranged between 0.431 and 0.972 with an average of 0.812 and from 0.782 to 0.949 with an average of 0.896, respectively. No significant linkage disequilibrium was observed after Bonferroni revision in any loci. The results show that the 15 polymorphic nuclear microsatellite markers can be used to study the population and conservation genetics of S. quinqueradiata in Korea. To ensure the success of artificial seedling production technology, genetic variations between the parent and offspring populations should be monitored, and inbreeding should be controlled.

유전적 다양성 분석을 활용한 꼬막의 원산지 판별

남보혜,동춘매,이미난,김은미,김영옥,노은수 한국패류학회 2024 The Korean Journal of Malacology Vol.40 No.1

This study aimed to develop a genetic analysis method to identify the origin of cockles (Tegilarca granosa). Initially, analysis of the mitochondrial COI gene revealed genetic variations among Korean, Chinese, and Japanese cockles. While distinguishing between Chinese and Japanese cockles was challenging, a specific haplotype (H1) found only in Korean cockles allowed researchers to infer their origin. Next, we analyzed the genetic characteristics of cockle populations using 19 microsatellite markers. This analysis validated the accuracy of assigning a cockle's origin and assessed the reliability of this method. The analysis revealed that three genetic loci (Teg02, Teg08, TMP18) displayed differing numbers of alternative alleles expressed between domestic populations and Chinese and Japanese populations, indicating genetic differentiation due to geographic separation. Using 16 microsatellite markers, the accuracy and reliability of origin determination achieved 89.6% and 86.6%, respectively. This shows there's enough information to distinguish between Korean, Chinese, and Japanese cockles. The results of this study demonstrate the usefulness of genetic analysis methods in determining the origin of cockles. This method is expected to contribute significantly to ensuring the safety and quality management of cockle distribution in the future. Additionally, there is a need for continued research to improve the accuracy and ability to differentiate origins through further development of microsatellite marker-based methods.

Four Members of Heat Shock Protein 70 Family in Korean Rose Bitterling (Rhodeus uyekii)

김정현,동춘매,김주란,안철민,백혜자,공희정 한국발생생물학회 2015 발생과 생식 Vol.19 No.3

Heat shock protein (HSP) 70, the highly conserved stress protein families, plays important roles in protecting cells against heat and other stresses in most animal species. In the present study, we identified and characterized four Hsp70 (RuHSP4, RuHSC70, RuHSP12A, RuGRP78) family proteins based on the expressed sequence tag (EST) analysis of the Korean rose bitterling R. uyekii cDNA library. The deduced RuHSP70 family has high amino acid identities of 72-99% with those of other species. Phylogenetic analysis revealed that RuHsp70 family clustered with fish groups (HSP4, HSC70, HSP12A, GRP78) proteins. Quantitative RT-PCR analysis showed the specific expression patterns of RuHsp70 family members in the early developmental stages and several tissues in Korean rose bitterling. The expression of 4 groups of Hsp70 family was detected in all tested tissue. Particularly, Hsp70 family of Korean rose bitterling is highly expressed in hepatopancreas and sexual gonad (testis and ovary). The expression of Hsp70 family was differentially regulated in accordance with early development stage of Rhodeus uyekii