• 대한의생명과학회지
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• 제24권4호
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• pp.292-304
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• 2018

Microbes is becoming increasingly forensic possibility as a consequence of advances in massive parallel sequencing (MPS) and bioinformatics. Human DNA typing is the best identifier, but it is not always possible to extract a full DNA profile namely its degradation and low copy number, and it may have limitations for identical twins. To overcome these unsatisfactory limitations, forensic potential for bacteria found in evidence could be used to differentiate individuals. Prokaryotic cells have a cell wall that better protects the bacterial nucleoid compared to the cell membrane of eukaryotic cells. Humans have an extremely diverse microbiome that may prove useful in determining human identity and may even be possible to link the microbes to the person responsible for them. Microbial composition within the human microbiome varies across individuals. Therefore, MPS of human microbiome could be used to identify biological samples from the different individuals, specifically for twins and other cases where standard DNA typing doses not provide satisfactory results due to degradation of human DNA. Microbial forensics is a new discipline combining forensic science and microbiology, which can not to replace current STR analysis methods used for human identification but to be complementary. Among the fields of microbial forensics, this paper will briefly describe information on the current status of microbiome research such as metagenomic code, salivary microbiome, pubic hair microbiome, microbes as indicators of body fluids, soils microbes as forensic indicator, and review microbial forensics as the feasibility of microbiome-based human identification.

• 한국자원식물학회지
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• 제23권6호
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• pp.526-534
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• 2010

Genomes of clusters of related eukaryotes are now being sequenced at an increasing rate. In this paper, we developed an accurate, low-cost method for annotation of gene prediction and exon-intron structure. The gene prediction was adapted for delta 1-pyrroline-5-carboxylate-synthetase (p5cs) gene from China wild-type of the halophytic Leymus chinensis (Trin.), naturally adapted to highly-alkali soils. Due to complex adaptive mechanisms in halophytes, more attentions are being paid on the regulatory elements of stress adaptation in halophytes. P5CS encodes delta 1-pyrroline-5-carboxylate-synthetase, a key regulatory enzyme involved in the biosynthesis of proline, that has direct correlation with proline accumulation in vivo and positive relationship with stress tolerance. Using analysis of reverse transcription-polymerase chain reaction (RT-PCR) and PCR, and direct sequencing, 1076 base pairs (bp) of cDNA in length and 2396 bp of genomic DNA in length were obtained from direct sequencing results. Through gene prediction and exon-intron structure verification, the full-length of cDNA sequence was divided into eight parts, with seven parts of intron insertion. The average lengths of determinated coding regions and non-coding regions were 154.17 bp and 188.57 bp, respectively. Nearly all splice sites displayed GT as the donor sites at the 5' end of intron region, and 71.43% displayed AG as the acceptor sites at the 3' end of intron region. We conclude that this method is a cost-effective way for obtaining an experimentally verified genome annotation.

• Parasites, Hosts and Diseases
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• 제50권4호
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• pp.391-394
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• 2012

The prevalence of liver and intestinal fluke infections was determined by surveying inhabitants of Hengxuan, Fusui, and Shanglin villages which were known to be endemic for liver flukes in Guangxi, China in May 2010. A total of 718 people were examined for helminth eggs by the Kato-Katz thick smear technique, ultrasonography, immunoaffinity chromatography, and DNA sequencing. The overall egg positive rate was found to be 59.6% (28.0-70.6%) that included mixed infections with liver and intestinal flukes. Cases showing higher than 20,000 eggs per gram of feces (EPG) were detected between 1.3% and 16.2%. Ultrasonographic findings exhibited overall 28.2% (72 of 255 cases) dilatation rate of the intrahepatic bile duct. Clonorchis sinensis infection was detected serologically in 88.3% (38 of 43 cases) among C. sinensis egg positive subjects by the immunoaffinity chromatography using a specific antigen for C. sinensis. For differential diagnosis of the liver and intestinal flukes, more precise PCR and nucleotide sequencing for copro-DNA were performed for 46 egg positive cases. Mixed infections with C. sinensis and Metagonimus yokogawai were detected in 8 of 46 egg positive cases, whereas 29 specimens were positive for Haplorchis taichui. Ultrasonographic findings and immunoaffinity chromatography results showed usefulness, even in a limited way, in figuring out of the liver fluke endemicity.

• Asian Pacific Journal of Cancer Prevention
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• 제15권21호
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• pp.9283-9289
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• 2014

Background: Acute lymphoblastic leukemia (ALL) is the most common cancer diagnosed in children and represents approximately 25% of cancer diagnoses among those younger than 15 years of age. Materials and Methods: This study investigated alterations in the displacement loop (d-loop) region of mitochondrial DNA (mtDNA) as a risk factor and diagnostic biomarker for early detection and diagnosis of acute lymphoblastic leukemia. Using mtDNA from 23 subjects diagnosed with acute lymphoblastic leukemia, the first 450 bp of the d-loop region were amplified and successfully sequenced. Results: This revealed 132 mutations at 25 positions in this region, with a mean of 6 alterations per subject. The d-loop alterations in mtDNA in subjects were all identified as single nucleotide polymorphisms in a homoplasmic distribution pattern. Mutant alleles were observed in all subjects with individual frequency rates of up to 95%. Thirteen mutant alleles in the d-loop region of mtDNA occurred with a high frequency. Novel alleles and locations were also identified in the d-loop of mtDNA as follows: 89 G insertions (40%), 95 G insertions (13%), 182 C/T substitutions (5%), 308 C insertions (19%), and 311 C insertions (80%). The findings of this study need to be replicated to be confirmed. Conclusions: Further investigation of the relationship between mutations in mitochondrial d-loop genes and incidence of acute lymphoblastic leukemia is recommended.

• Clinical and Experimental Pediatrics
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• 제63권6호
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• pp.195-202
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• 2020

Developments in next-generation sequencing (NGS) techogies have assisted in clarifying the diagnosis and treatment of developmental delay/intellectual disability (DD/ID) via molecular genetic testing. Advances in DNA sequencing technology have not only allowed the evolution of targeted panels but also, and more currently enabled genome-wide analyses to progress from research era to clinical practice. Broad acceptance of accuracy-guided targeted gene panel, whole-exome sequencing (WES), and whole-genome sequencing (WGS) for DD/ID need prospective analyses of the increasing cost-effectiveness versus conventional genetic testing. Choosing the appropriate sequencing method requires individual planning. Data are required to guide best-practice recommendations for genomic testing, regarding various clinical phenotypes in an etiologic approach. Targeted panel testing may be recommended as a firsttier testing approach for children with DD/ID. Family-based trio testing by WES/WGS can be used as a second test for DD/ID in undiagnosed children who previously tested negative on a targeted panel. The role of NGS in molecular diagnostics, treatment, prediction of prognosis will continue to increase further in the coming years. Given the rapid pace of changes in the past 10 years, all medical providers should be aware of the changes in the transformative genetics field.

• 한국수정란이식학회지
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• 제10권1호
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• pp.91-100
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• 1995

3.3kb 웅성특이 DNA(pEM39 plasmid DNA)가 성 특이 DNA 검색자로 활용되어질 수 있는가를 확인하기 위하여 구조적인 분석을 Southern blotting, DNA sequencing과 computer program 분석을 통하여 실시하였다. 전체 3.3kb에서 유래된 약 1kb 단위의 단편을 이용하여 표지된 짧은 DNA probe들은 Southern blot 분석에서 웅성특이성을 나타내었다. McGraw와 Jeon의 sequence에 대한 유사성 비교 자료로부터 여러 부분의 conserved region을 찾아내고 이것을 기초로 하여 5개의 primer set들을 선발하였다. Conserved region에 존재하면서 computer program에 의해서 선발되어진 PMS1과 2의 primer set가 최종적으로 PCR 분석을 위하여 선정되었다. 이 primer set를 사용한 PCR 분석에서, 1ng부터 10pg까지의 웅성 genomic DNA에서 PCR 산물을 얻을 수 있었으며, 자성의 경우는 어떠한 산물도 찾을 수 없었다. PCR에 이용할 수정란의 시료는 2 세포기의 수정란에서 얻었으며 순수 분리된 genomic DNA에서 확립된 조건에서 PCR을 수행하였다. 8개의 수정란을 분석한 결과 4개의 웅성과 4개의 자성 수정란을 확인하였다. 이러한 결과는 선정된 primer set가 돼지 수정란의 성을 조기 감별하는데 효율적인 DNA probe로 사용될 수 있다는 것을 암시한다.

• 한국축산식품학회지
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• 제37권4호
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• pp.535-541
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• 2017

This study investigated the psychrotrophic bacteria isolated from chicken meat to characterize their microbial composition during refrigerated storage. The bacterial community was identified by the Illumina MiSeq method based on bacterial DNA extracted from spoiled chicken meat. Molecular identification of the isolated psychrotrophic bacteria was carried out using 16S rDNA sequencing and their putrefactive potential was investigated by the growth at low temperature as well as their proteolytic activities in chicken meat. From the Illumina sequencing, a total of 187,671 reads were obtained from 12 chicken samples. Regardless of the type of chicken meat (i.e., whole meat and chicken breast) and storage temperatures ($4^{\circ}C$ and $10^{\circ}C$), Pseudomonas weihenstephanensis and Pseudomonas congelans were the most prominent bacterial species. Serratia spp. and Acinetobacter spp. were prominent in chicken breast and whole chicken meat, respectively. The 118 isolated strains of psychrotrophic bacteria comprised Pseudomonas spp. (58.48%), Serratia spp. (10.17%), and Morganella spp. (6.78%). All isolates grew well at $10^{\circ}C$ and they induced different proteolytic activities depending on the species and strains. Parallel analysis of the next generation sequencing and culture dependent approach provides in-depth information on the biodiversity of the spoilage microbiota in chicken meat. Further study is needed to develop better preservation methods against these spoilage bacteria.

• Clinical and Experimental Pediatrics
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• 제45권10호
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• pp.1263-1272
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• 2002

목 적 : 레트증후군이란 1966년 Andreas Rett에 의해 처음 기술되었으며 생후 6개월에서 18개월 정도까지 비교적 정상 발달을 한 후 두위 발달의 감소와 함께 습득했던 인지 및 운동능력의 상실, 언어기능의 상실, 그리고 특징적인 손동작(상동행동)을 보이는 X 염색체 우성유전으로 생각되는 질환이다. 1999년 미국에서 그 결함 유전자인 MECP2 유전자가 Xq28에서 밝혀졌으나 우리 나라에서는 이 질환에 대한 전반적 이해가 부족하며 유전자 연구는 거의 전무한 상태이다. 이에 저자들은 우리 나라의 Rett 증후군 환아에서 MECP2 유전자의 결함이 어느 정도 나타나는지를 알고자 이 연구를 시행하였다. 방 법 : 2000년 2월부터 2001년 3월까지 본원을 방문한 Rett 증후군 환아 34례의 말초혈액을 EDTA (Ethylenediaminetetraacetic acid) 처리하여 5 cc 채취하여 냉동 보관한 후 해빙하여 DNA 표본을 추출했다. DNA의 추출은 E.Z.N.A blood DNA kit을 사용하였다. MECP2 유전자 네 종류의 axon은 PCR을 이용해 증폭하였고 primer sequence는 1999년 Amir 등에 의해 디자인 된 것(AF030876)을 사용하였다. ABI 377 DNA sequencer와 ABI PRISM dye cycle sequencing reaction kit을 사용하여 DNA sequencing을 시행하였고 우리 나라에서 최초로 발견된 유전자 변이에 대해서는 RFLP 분석을 통해 확인하였다. 결 과 : 1) MECP2 유전자의 변이를 보인 환아는 23례로 67.6%에서 관찰되었다. 2) 9종의 missense mutation과 3종의 nonsense mutation을 합해 총 12종의 유전자 변이가 관찰되었다. 3) 이들 돌연 변이 중 L100V, G161E, 그리고 T311M은 전세계적으로 우리 나라에서 처음 발견된 변이이다. 4) 23례의 유전자 변이는 대부분(78.3%) MBD와 TRD라는 기능영역에서 발견되었다. 5) 본 연구에서 T158M, R270X, 그리고 R306C가 자주 나타나는 유전자 변이였다. 결 론 : 우리 나라의 Rett 증후군 환아에서도 MECP2 유전자의 변이는 비교적 흔히 관찰되어 MECP2 유전자의 이상이 Rett 증후군을 유발하는 주 유전자 이상임을 확인하였고 Rett 증후군 환아의 확진을 위해 MECP2 유전자 검사가 필요할 것으로 사료된다.

• 생태와환경
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• 제54권3호
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• pp.247-256
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• 2021

동물플랑크톤이 식물플랑크톤을 선택적으로 섭식하는 특성에 대한 이해는 수생태계 먹이사슬 내의 물질 이동에 중요하다. 하지만 해부를 통한 위내용물 추출 방법은 소형 요각류를 대상으로 적용하기에는 적절하지 않고, 유전자가 유실되거나 위내용물이 아닌 개체 외부의 유전자로 인해 오염될 가능성이 존재한다. 본 연구에서 호 내 식물 플랑크톤 조성 및 기타 환경이 상이한 두 지점을 선정하여 모든 지점에서 지속적으로 출현하는 기수성 요각류인 Sinocalanus tenellus를 대상으로 위내용물의 유전자 분석을 수행하였다. 요각류 개체 외부의 DNA를 제거하는 데 2.5%로 희석한 시판용 표백제(차아염소산나트륨 5.4%)에 2분간 처리하여 증류수로 2회 세척한 뒤 유전자를 추출하였다. 추출된 유전자는 23S rRNA을 증폭하여 서열분석을 실시하였다. Capillary sequencing 분석 결과, 원수와 처리수 및 요각류 위내용물에서 다양한 분류군(규조강, 녹조강, 남조강, 와편모조강, 은편모조강, 황갈조강)에 속하는 식물플랑크톤이 검출되었으며, 새만금호 내 시공간 차이에 따라 상이한 경향을 보였다. 현미경을 이용하여 동정한 식물플랑크톤 군집 조성의 경우 규조강이 우점한 반면, 동일한 원수의 유전자 분석(capillary sequencing) 결과에서는 주로 녹조강, 남조강 및 와편모조강이 우점하여 다소 상반된 경향을 나타냈다. 본 연구에서 적용한 위내용물 분석에 특화된 외부 유전자 제거 전처리 방법은 농도와 처리시간 조절 등의 응용방법에 따라 다양한 동물플랑크톤 분류군에 적용이 가능할 것으로 사료된다.

• Asian-Australasian Journal of Animal Sciences
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• 제22권8호
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• pp.1091-1101
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• 2009

In an attempt to understand the biochemical mechanism for the synthesis of the anabolic steroid, 19-nortestosterone, produced by prepubertal boar testes and its physiological role, normalized complementary DNA (cDNA) from boar testes was generated. DNA sequencing of 2,016 randomly selected clones yielded 794,116 base pairs of high quality nucleotide sequence. Computer-assisted assembly of the nucleotide sequence of each clone resulted in 423 contigs and 403 singletons including several genes for steroidogenic enzymes and molecules related to steroid metabolism. Analysis of gene expression pattern by use of the presently-fabricated cDNA microarray identified a number of genes that were differentially expressed during the postnatal development period in boar testes. Two genes of unknown function were identified to be highly expressed in the testis of 2-weeks-old neonatal boar. In addition, the sequencing of open reading frames of these genes revealed their homology with human alpha hemoglobin and Homo sapiens hypothetical LOC643669, transcript variant 1. Moreover, the transcripts of these genes were also detected in porcine muscle and adipocytes, in addition to Leydig cells of pigs.