• 한국미생물·생명공학회지
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• 제23권2호
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• pp.145-149
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• 1995

For the purpose of developing a new biodegradable detergent, we have isolated a gene encoding wide-range temperature applicable alkaline protease from Xanthomonas sp. YL-37 (Lee et al., 1994, Kor. J. Appl. Microbiol. Biotechnol.). An alkaline protease gene was isolated from the gene bank that was prepared from the chromosomal DNA of Xanthomonas sp. YL-37. From the results of agarose gel electrophoresis and a restriction enzyme mapping, a 2.7 kb DNA fragment containing the alkaline protease gene was inserted in the plasmid pUC9. Extracellular activity of a clone having alkaline protease gene was detected on SDS-polyacrylamide gel with activity staining assay. The molecular weight of alkaline protease was determined to be about 64 kDa from 11% SDS-PAGE analysis. Alkaline protease activity, produced from E. coli which harboring the plasmid, showed no difference at reaction temperature 20, 30 and 40$\circ$C, respectively. This result showed that alkaline protease produced from E. coli harboring the plasmid was apparently the same as that of Xanthomonas sp. YL-37.

• 한국연초학회지
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• 제21권1호
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• pp.57-63
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• 1999

Pokeweed antiviral protein II (PAP-II) encoding cDNA was synthesized by reverse-transcriptase polymerase chain reaction (RT-PCR) from Phytolacca american a leaf. The PAP-II cDNA fragment of 974bp was subcloned to pBluescript II SK- SmaI site and the inserted PAP-II cDNA fragment was sequenced by dideoxy sequencing method. The number of nucleotides of PAP-II cDNA coding region containing start and stop codon was 933bp. To develop a virus-resistant tobacco plant, PAP-II cDNA fragment was inserted to pKGT101B and the insertion of PAP-II cDNA fragment was confirmed by restriction enzyme analysis and colony PCR.

• Advances in Traditional Medicine
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• 제7권4호
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• pp.357-362
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• 2007

Cytochrome P450 2C19 (CYP2C19) is a clinically important enzyme involved in the metabolism of therapeutic drugs, including (S)-mephenytoin, omeprazole, proguanil, and diazepam. Individuals are characterized as either extensive metabolizers (EM) or poor metabolizers (PM) on the basis of CYP2C19 enzyme activity. The PM phenotype occurs in 2-5% of Caucasians, but in 18-23% of Asians. To clarify the association between CYP2C19 polymorphisms and gastric cancer in Koreans, we investigated CYP2C19 genotypes ($CYP2C19^*1,\;{^*2},\;and\;^*3$) in 109 patients with gastric cancer and 211 controls. Normal ($CYP2C19^*1$) and defective alleles were detected with polymerase chain reaction/restriction enzyme analysis. CYP2C19 has three hereditary genotypes: homozygous EM, with high enzymatic activity; heterozygous EM, with moderate enzymatic activity; and PM, with no enzyme activity. We found that CYP2C19 heterozygous EM is more closely associated with gastric cancer than is homozygous EM. Because the CYP2C19 genotype varies in Koreans, a genotyping test is desirable to prevent gastropathy recurrence in patients before their doses of omeprazole are reduced during maintenance therapy.

• Asian-Australasian Journal of Animal Sciences
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• 제16권5호
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• pp.650-654
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• 2003

The accuracy of Polymerase chain reaction (PCR) assay in sexing of sheep embryos was assessed in this study. A total of 174 ovine embryos produced in vitro at different stages of development (2, 4-8 cell stages, morula and blastocyst) were sexed. The universal primers (P1-5EZ and P2-3EZ) used in this assay amplified ZFY/ZFX-specific sequences and yielded a 445 bp fragment in both sexes. Restriction enzyme analysis of ZFY/ZFX-amplified fragments with Sac I exhibited polymorphism between sexes, three and two fragments in males and in females, respectively. For verification of accuracy, blood samples of known sex were utilized as positive controls in each test. The mean percentages of sex identification by this method at 2 cell, 4-8 cell, morula and blastocyst were $73.00{\pm}5.72$, $89.77{\pm}3.79$, $3.33{\pm}8.08$ and $79.6{\pm}9.09$, espectively with the over all male to female ratio of 1:0.87. It is concluded that the ZFY/ZFX based method is highly reliable for the sexing of sheep embryos.

• 농업과학연구
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• 제47권3호
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• pp.673-681
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• 2020

Human enteric Adenovirus 41 (HueAdV-41) is a major waterborne virus that causes human gastroenteritis and is classified as a viral group I double-strand DNA virus, Adenoviridae. HueAdV-41 has been detected with the polymerase chain reaction (PCR) in various samples such as ground water. However, the PCR-based diagnostic method has problems such as reaction time, sensitivity, and specificity. Thus, the loop-mediated isothermal amplification (LAMP) assay has emerged as an excellent method for field applications. In this study, we developed a LAMP system that can rapidly detect HueAdV-41 with high specificity and sensitivity. HueAdV-41 specific LAMP primer sets were tested through a specific, non-specific selection and sensitivity test for three prepared LAMP primer sets, of which only one primer set and optimum reaction temperature were selected. The developed LAMP primer set condition was confirmed as 63℃, and the sensitivity was 1 copy. In addition, to confirm the system, a LAMP positive reaction was developed with the restriction enzyme Taq I (T/GCC). The developed method in this study was more specific, rapid (typically within 2 - 3 hours), and highly sensitive than that of the conventional PCR method. To evaluate and verify the developed LAMP assay, an artificial infection test was done with five cDNAs from groundwater samples, and the results were compared to those of the conventional PCR method. We expect the developed LAMP primer set will be used to diagnose HueAdV-41 from various samples.

• 한국양식학회지
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• 제6권3호
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• pp.221-233
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• 1993

범가자미, Verasper variegatus에 대한 유전학적 동정을 위하여 세포 크기, DNA함량, 염색체수 및 핵형분석 등의 세포유전학적 조사와 PCR 기법을 이용한 mtDNA 125 ribosomal RNA gene의 분석을 실시하였다. 본 종의 적혈구와 핵의 평균 부피는 각각 $211.10{\mu}m^3$와 $23.03{\mu}m^3$였으며, haploid DNA content는 0.79 pg/cell로서 잉어의 $46.5\%$, 포유류의 $22.6\%$로 나타났다. 염색체 수는 46개로 모두 acrocentric 염색체로 구성되어 있었으며, heteromorphic한 성 염색체는 관찰되지 않았다. PCR 기법을 이용하여 증폭된 범가자미 mtDNA의 12S rRNA gene segment는 대략 390bp로 나타났고, 12S rRNA gene의 PCR product를 제한 효소로 처리 결과, Ava I, Mae II, Sma I, Xba I는 1개의 restriction site가, Mae I는 2개의 restriction site가 관찰되었다. 범가자미 mtDNA의 12S rRNA gene segment의 염기 서열을 인간과 차넬메기와 비교한 결과, identity가 차넬메기 와는 $81.8\%$, 인간과는 $67.7\%$였다.

• Korean Chemical Engineering Research
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• 제49권2호
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• pp.129-136
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• 2011

SMB 공정은 주로 4개의 구역으로 나뉘어지는 다수의 크로마토그래피 컬럼으로 구성된다. 이러한 특성은 회분식 크로마토그래피 공정보다 우수한 이성분계 물질의 연속적인 분리를 구현한다. SMB는 회분식 크로마토그래피에 비해 연속성 및 높은 생산성과 순도로 목적물질을 분리해 낼 수 있는 장점을 갖는다. 경제적이며 효율적인 공정의 운용을 위해 반응과 회수를 결합시키는 연구가 보고되고 있으며, 이와 같은 연구 중 SMBR은 연속분리공정인 SMB와 반응기가 결합된 공정이다. 다양한 반응을 적용한 SMBR에 대해 많은 연구가 진행되고 있으며, 촉매반응, 효소반응, 이온 교환 수지를 통한 화학반응이 주를 이루고 있다. 초기의 SMBR은 촉매를 사용한 고정층의 형태이며, 유동성 효소를 사용하는 SMBR, 고정화 효소를 사용하는 SMBR, 반응구역과 흡착구역이 분리되어 있는 SMBR순으로 발전하였다. 공정 설계에 있어서 필수적인 모델링 및 최적화를 위하여 대류현상만을 고려한 간단한 기법이 있지만, 실제 물질거동을 설명하기 위해서는 축 방향 분산이나 물질전달 저항을 고려한 복잡한 해석을 필요로 한다. SMBR같이 반응과 분리가 결합된 공정의 경우 설비의 간소화를 통한 시설비용의 축소뿐 아니라 가역반응평형의 극복을 통해 물질의 순도와 수율을 향상시킬 수 있는 장점이 있다.

• 한국가축번식학회지
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• 제23권2호
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• pp.159-163
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• 1999

최근 분자생물학적 기법의 발달은 유전병에 대한 조기진단과 경제형질 관련 유전자에 대한 동정과 분석을 가능하게 하였다. 본 연구에서는 내분비학적 실험을 목적으로 채취된 돼지의 혈액으로부터 혈청을 추출한 후, 응고된 혈액과 이를 건조시킨 혈액으로부터 유전 분석을 실시하기 위해 genomic DNA를 추출하였다. 또한, 추출된 DNA를 이용하여 에스트로겐 수용체 유전자에 대한 PCR을 수행하여, 그 산물을 적절한 제한 효소를 처리하여 유전자 다형 현상을 관찰할 수 있었다. 따라서 본 연구에서는 돼지의 내분비 물질 분석 실험의 부산물인 응고ㆍ건조된 혈액을 사용하여 경제적이면서 신속하게 분석하고자 하는 개체의 유전자형을 밝힐 수 있으며, 내분비학적ㆍ분자유전학적 분석방법을 동시에 수행함으로써 내분비 물질 발현과 유전자형간의 관계를 구명할 수 있는 가능성을 제시하였다.

• Journal of Genetic Medicine
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• 제2권1호
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• pp.17-22
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• 1998

The Lesch-Nyhan syndrome which is caused by the deficiency of hypoxanthine guanine phosphoribosyltransferase is an X-linked recessive disorder characterized by hyperuricemia, choreoathetosis, mental retardation and compulsive self-injurious behavior. Clinical management of the patients with the Lesch-Nyhan syndrome is frustrating and requires burdensome medical treatment since it cripples the patient and shortens the life span by progression of neurological symptoms, but there are no cures or measures for relieving relentless natural course of the disease yet. Therefore, prenatal diagnosis of the affected fetus is important in genetic counselling for the family at high risk. In this study, four different mutations in the HPRT gene of four probands have been identified in four unrelated families; K215X, Q109X, nt.631 ${\Delta}A$, and nt.289 ${\Delta}GT$. Two mutations among them altered restriction enzyme sites; SpeI for Q109X and MaeI for nt.289 ${\Delta}GT$. Based on their molecular defects, prenatal diagnoses of 3 the fetuses were successfully made between ninth and eleventh week of gestation by polymerase chain reaction (PCR), restriction digestion and DNA sequencing using cDNA obtained from chorionic villus samples (CVS). We predicted the outcome of all fetuses prenatally. Among the three fetuses two were male and one was female according to the identification made by PCR amplification of the sex determining region of the Y chromosome(SRY) gene. Each carried a wild type allele for the corresponding mutant allele. They were also tested postnatally for the mutations to be unaffected.

• The Plant Pathology Journal
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• 제18권2호
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• pp.85-92
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• 2002

A new isolate of Cucumber mosaic virus (CMV), identified as Li-CMV was isolated from a diseased Korean native lily (Lithium tsingtauense Gilg). Biological and serological properties of Li-CMV were characterized, and reverse transcription-polymerase chain reaction (RT-PCR) analysis, restriction enzyme profiling of RT-PCR products, and nucleotide sequence analysis of RNA3 of the virus were performed in this study. Remarkable differences in symptoms between Li-CMV and ordinary CMV strains were found in tobacco plants and Datura stramonium. Li-CMV-infected tobacco plants (cv. Xanthi-nc and cv. Samsun) induced chlorotic ringspots on uninoculated upper leaves, and the symptom expression was delayed or faint whereas, ordinary CMV strains induced green mosaic symptoms on the plant. Systemic infections were observed on Nicotiana benthamiana with severe mosaic symptom. Restriction mapping analysis of RT-PCR products using MspI showed that Li-CMV belonged to CMV subgroup I. A full-length CDNA copy of RNA3 for the virus was amplified by RT-PCR, cloned, and its complete nucleotide sequence was determined. The RNA3 of Li-CMV was 2, 232 nucleotides long, and consisted of two open reading frames of 843 and 657 bases encoding 3a protein (movement protein) and coat protein, respectively. Results of this study indicate that Li-CMV is a novel strain and belongs to subgroup I of CMV in the genus Cucumovirus.