• 미생물학회지
• /
• 제13권3호
• /
• pp.109-115
• /
• 1975

DNA's were extracted from Saccharomyces cerevisiae and Mycobacterium phlei and the damaging cell walls of these microoragnisms were examined under an electron microscope in the extraction process in which a number of physico-chemical tratments of cells was involved. While the DNA was easily extracted from S. cerevisiae using conventional meylelded very little DNA, of M. phlei was extremely difficult to isolate and yielded very little DNA, applying various methods of isolation published earlier. When the cell walls of S. cerevisiae were examined with the electron microscope, they were not yet damaged even after the cells were treated with sodium lauryl sulfate(SLS) and ethylene diamine tetracetic acid(EDTA), but they were completely destroyed by the treatment of sodium perchlorate followed by the addition of chloroform and a vigorous agitation. Oozing cytoplasm through the broken cell walls was also observed. In the extraction of DNA from M.phlei, the pronase was not effective at the aerobic environment of the sample. When phenol was applied at the last step of DNA isolation, an extreme damage mass yielding little DNA into the solution. Unlike the cells of S.cerevisiae.M.phlei cells showed a tendency of aggregation, thus the destruction of cell walls by sodium hydroxide was seen only on the walls of peripheral cells in the aggregated mass, leaving the walls of the inner cells undamaged.

• ALGAE
• /
• 제28권4호
• /
• pp.371-378
• /
• 2013

Water and methanolic extracts of Ecklonia cava, a marine brown alga, were prepared by ultrasonic extraction (UE) and conventional extraction (CE) methods. The radical-scavenging activity and the inhibitory effects against hydrogen peroxide ($H_2O_2$)-induced DNA damage of the extracts were investigated. All extracts prepared by CE exhibited higher total polyphenolic content than that in the extracts prepared by UE. Extraction yield and total phenolic content increased as the UE time increased. The radical-scavenging activities increased as the UE time increased. All extracts prepared by CE exhibited higher 1,1-diphenyl-2-pricrylhydrazyl (DPPH) and hydroxyl radical-scavenging activities than did those prepared by UE. Extracts prepared by UE showed stronger scavenging activities on alkyl radical and $H_2O_2$ than those prepared by CE did. Methanolic extract with UE 12 h (100MEU-12h) and methanolic extract with CE 24 h (100MEC-24h) were selected and evaluated by comet assay for their inhibitory effect against $H_2O_2$-induced DNA damage. 100MEU-12h showed slightly greater protective effect against $H_2O_2$-induced DNA damage than 100MEC-24h. Thus, UE can be effectively used as a seaweed extraction technique, and there is potential for scale-up of the extraction process.

• 한국자원식물학회지
• /
• 제14권3호
• /
• pp.235-240
• /
• 2001

Polyphenol 화합물이 다량 함유된 식물종의 유연관계 분석이나 형질전환 유전자 확인 등을 위해 PCR을 이용할 경우 다량의 재료로부터 신속 간편하게 분리한 DNA를 이용할 수 있는 조건을 설정 하였다. 폴리페놀 함량이 높은 포도, 사과, 복분자와 같은 과수류에서 간편법에 의해 추출된 DNA를 이용한 PCR 반응액에 2%의 BLOTTO를 첨가함으로서 DNA의 재현적 증폭이 가능하였다. 간편 추출 DNA를 이용한 PCR에서 의 BLOTTO효과는 primer, 품종, 식물종에 관계없이 일반적으로 발현되었다. 상추의 형질전환 유전자 검색을 위한 PCR에서 도 BLOTTO 효과가 확인되었다. 따라서 PCR 반응액에 2% BLOTTO를 첨가하면 간편 법 에 의해 추출된 polyphenol 화합물 고함유 식물종의 DNA를 이용하여서도 PCR에 의한 유전배경 및 특정 유전자의 대량 신속 분석이 가능할 것이다.

• 한국식품위생안전성학회지
• /
• 제36권1호
• /
• pp.93-99
• /
• 2021

본 연구에서는 PCR법을 이용하여 농산물 중 enterotoxin 생성 Clostridium perfringens를 신속 분석할 수 있도록 전 처리법을 확립하고자 수행하였다. 이를 위하여 C. perfringens 포자를 상추, 토마토, 고추, 들깻잎에 102, 103, 104, 105, 106, 107 spore/g 농도로 포자를 접종하였다. 포자가 접종된 농산물들은 pulsifier, stomacher, sonicator로 처리하고 boiling법 혹은 상용화 된 kit로 DNA를 추출한 후 PCR법을 수행하고 검출한계를 비교하였다. 그 결과, 3가지 전처리법에 있어서는 pulsifier가, DNA 추출에 있어서는 상용화된 DNA 추출 kit를 활용하는 것이 농산물 중 C. perfringens의 검출한계를 10-100배 낮출 수 있었다. 특히 들깻잎, 방울토마토처럼 전처리 방법에 따른 탁도의 변화가 큰 농산물은 전처리법과 DNA 추출법이 PCR 반응에 미치는 영향이 큰 것으로 나타났다. 따라서 본 연구 결과를 통해 볼 때 PCR법을 이용한 농산물 중 C. perfringens를 검출하는데 있어 검출감도를 높이기 위해서는 pulsifier를 이용하여 전처리하고 상용화된 DNA 추출 kit를 사용하는 것이 적절한 것으로 판단된다.

• 보존과학연구
• /
• 통권22호
• /
• pp.27-40
• /
• 2001

DNA typing is often used to determine identity from human remains. Recently, the molecular biological analysis of ancient deposits has become possible since methods for the recovery of DNA conserved in bones or teeth from archaeological remains have been developed. In the field of archaeology, one of the most promising approaches is to identify the individuals present in a mass burial site. We performed nuclear DNA typing and mitochondrial DNA sequencing analysis based on PCR from a Korea ancient human remain excavated from Sa-chon Nuk-island and civilian access controlline(CACL). A femur bone were collected and successfully subjected to DNA extraction, quantification, PCR amplification, and subsequently typed for several shot tandem repeat(STR)loci. 4 types of STR systems used in this study were CTT multiplex(CSF1PO, TPOX, TH01), FFv multiplex(F13A01, FESFPS, vWA), Silver STRⅢ multiplex(D16S539, D7S820, D13S317), and amelogenin for sex determination. This studies are primarily concerned with the extraction, amplification, and DNA typing of ancient human bone DNA samples. Also, it is suggestive of importance about closely relationship between both fields of archaeology and molecular biology.

• Journal of Oral Medicine and Pain
• /
• 제20권2호
• /
• pp.497-513
• /
• 1995

Human genomic Deoxyribonucleic acid(DNA) was extracted from teeth by boiling, salting-out, phenol, boiling-phenol methods. The author compared DNA concentration and its purity, the accuracy of sex determination and the results of the D1S80 locus detection among above 4 methods. The following results were obtained : 1. DNA concentration was the highest in pulp with salting-out method and DNA purity was higher in pulp with salting-out and phenol methods than other 2 methods. 2. Sex determination was possible using of the pulp and the dentin of the teeth with four methods but, it was impossible in the enamel and some pulp with boiling method. 3. Amplification of D1S80 locus occurred from pulp and dentin with salting-out, phenol, and boiling-phenol methods. 4. There are no differences among the amplification of X-Y homologus amelogenin gene by application of 4 methods and salting-out, phenol methods efficiently makes available to amplification of D1S80 locus. From the investigation DNA extraction, sex determination, amplification of D1S80 locus was successfully accomplished with salting-out, phenol, boiling-phenol methods Therefore above 3 methods are available and applicable as forensic odontology for individual identification.

• 한국식품과학회지
• /
• 제48권4호
• /
• pp.335-340
• /
• 2016

본 연구에서는 다양한 식품으로부터 식중독 세균의 DNA를 추출하는 효율을 비교 하였다. 사용된 DNA 추출 방법은 TaKaRa Kit를 이용하는 방법, PrepMan reagent를 이용하는 방법, boiling method, PEG를 이용한 alkaline method가 사용되었다. 비용절감이나 시간절약 면에서 boiling method나 PrepMan method도 고려할 수 있지만, column kit를 이용하는 TaKaRa kit가 효율적이라고 판단되었다. 또한 정성 시험법에서 적은 양의 균을 검출하기 위해 증균배양을 거치게 되는데 이때 사용되는 증균배지의 성분이 DNA 추출 후에도 잔류하여 saline과 비교하였을 때 DNA 추출효율이 낮은 결과를 나타내었다. 따라서 증균배지의 성분을 제거한 뒤 DNA를 추출하는 것이 PCR의 효율을 높일 수 있을 것으로 예상된다. 정량 시험법에서는 증균과정을 거치지 않아 균을 검출할 때 DNA의 추출 효율이 중요하기 때문에 DNA 추출 효율을 높이기 위한 가장 좋은 버퍼를 선정하고자 하였다. 그 결과 E. coli O157:H7에서는 saline이, S. aureus에서는 증류수를 버퍼로 사용했을 때 DNA 추출 효율이 가장 높은 것으로 나타났다.

• Fisheries and Aquatic Sciences
• /
• 제25권5호
• /
• pp.264-275
• /
• 2022

A non-destructive environmental DNA protocol for the genetic analysis of sea cucumber (Apostichopus japonicus) resources DNA was established. Among the several commercial DNA extraction kits, the DNeasy^® Plant Mini Kit was selected as the best choice to obtain the high-quality genomic DNAs from the mucous sea cucumber. As the temperature and incubation time increased, the amount of extracted environmental DNA was also large, but it was judged that the increased amount did not affect as much as 2-3 times. Therefore, these conditions were not considered to be the main factors to consider in actual environmental DNA extraction. However, the amount of seawater relative to the size of the sample was judged as a major consideration, and a sufficient amount of environmental DNA for analysis was secured when stored within 1 min while stirring the volume of seawater corresponding to the total sea cucumber weight (g). In securing the environmental DNA of sea cucumbers, the mortality rate of sea cucumbers in all experiments was 0, and it was judged that the effects of sea cucumbers were not significant through this treatment. Through the results of this study, sea cucumber DNA research, which has been conducted in a destructive method, can be conducted non-destructively through environmental DNA analysis. Through this study, we have secured a standard protocol that can successfully extract the sea cucumber DNA through environmental DNA. It is not only excellent in terms of time and cost of traditional DNA analysis method currently used, but it is completely non-destructive in the ecosystem of the survey area. It is believed that the system can be transformed in a way that does not affect it. However, it is thought that various standard protocols should be established considering the characteristics of each type.

• Journal of Genetic Medicine
• /
• 제7권1호
• /
• pp.53-58
• /
• 2010

목 적: 모체 혈장으로부터 가장 효과적으로 세포 유리 DNA(cell free DNA, cf-DNA)를 추출하는 방법을 찾기 위해 우리는 viral DNA 추출 방법과 일반 혈액DNA 추출 방법을 이용하여 비침습적 임신 초기 태아 성별 확인 결과를 비교하였다. 대상 및 방법: 임신 초기 44명의 임산부로부터 모여진 모체 혈장을 통한 전향적 연구가 구성되었다. Cf-DNA는 viral DNA 추출 방법과 일반 혈액 DNA 추출 방법을 이용하여 각각 추출되었다. 정량 형광-중합효소 연쇄 반응(QF-PCR)을 이용하여SRY 와AMXY 유전자를 검출하였다. QF-PCR의 진단 정확도는 최종 분만 기록을 토대로 결정하였다. 결 과: 전체 44명의 여성이 실험에 참여하였지만, 최종 분만 기록은 단지 36명의 여성에서 획득하였다. 이들 중 16명은 남아를 20명은 여아를 임신하였다. 두 추출 방법에서 태아 성별의 진단적 정확도는 일반 혈액 DNA 추출 방법에 경우 63.9% (23/26)였으며 viral DNA 추출 방법에 경우 97.2% (35/36) 였다. 결 론: QF-PCR을 이용한 비침습적 임신초기 태아 성별 확인에 있어 viral kit를 사용하는 것이 높은 진단적 정확도를 이끌 수 있을 것으로 사료된다.

• 대한약학회:학술대회논문집
• /
• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.1
• /
• pp.290.1-290.1
• /
• 2003

Oxidative DNA damage has been associated with many disease. Quantation of DNA adducts is considered to be a useful biomarker of oxidative DNA damage because its formation can also be induced by oxidative stress. Extensive efforts have been taken to identify the analytical methods for minimizing the artifactual formation of oxidative DNA damage. We have done direct analysis of DNA adducts using LC/ESI-MS without urine sample extraction. (omitted)