• 한국양식학회지
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• 제12권2호
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• pp.91-100
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• 1999

The complete nucleotide sequence of olive flounder (Paralichthys olivaceus) hsp70-related rDNA was determined by RT- and RACE-PCR methods. A full-length of hsp70-related cDNA has an open reading frame of 1.95 kb encoding 650 amino acids with a calculated molecular weight of 71.1 kD. A corresponding hsp70-related protein contains a number of conserved elements including an ATP-binding domain, a nuclear localization signal and the carboxyl terminal motif, EEVD, which may have a role in chaperone function. Comparison of nucleotide and predicted amino acid sequence between olive flounder hsp70-related gene and hsp/hsc70 genes of other species revealed a high similarity with the cognate form of these genes. These results indicated that we recovered likely to be a olive flounder cognate hsc70 gene.

• Journal of Microbiology and Biotechnology
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• 제13권2호
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• pp.230-235
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• 2003

A 1.25 kb DNA fragment including the lscR gene, which encodes a levansucrase of Rahnella aquatilis ATCC 15552, was subcloned into a high-expression vector, pET-29b, and the recombinant enzyme was overexpressed in Escherichia coli. Most of the levansucrase activity was detected in the cytoplasmic fraction after induction with isopropyl ${\beta}-D-thiogalactoside$. The recombinant enzyme with a tag of six histidine residues at the C-terminus was purified 146-fold by affinity and gel-filtration chromatographies. The molecular mass of the purified LscR was approx. 49 kDa as determined by SDS-PAGE. The optimum pH and temperature of this enzyme for levan formation was pH 6.0 and $30^{\circ}C$, respectively. The optimum substrate concentration for levan formation was 300 mM sucrose. Levan formation was increased by the increase of the enzyme concentrations. Maxium yield of levan formation at optimum substrate concentration, pH, and temperature after 24 h of reaction was approximately 80%.

• Ocean Science Journal
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• 제40권3호
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• pp.155-166
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• 2005

New PCR primers (N=18) were designed for the isolation of complete SSU to LSU rDNA sequences from the dinoflagellate Alexandrium tamarense. Standard PCR, employing each primer set selected for amplifications of less than 1.5 kb, successfully amplified the expected rDNA regions of A. tamarense (Korean isolate, HY970328M). Complete SSU, LSU rDNAs and ITS sequences, including 5.8S rDNA, were recorded at 1,800 bp, 520 bp and 3,393 bp, respectively. The LSU rDNA sequence was the first report in Alexandrium genus. No intron was found in the LSU rRNA coding region. Twelve D-domains within the LSU rDNA were put together into 1,879 bp (44.4% G+C), and cores into 1514 bp (42.8% G+C). The core sequence was significantly different (0.0867 of genetic distance, 91% sequence similarity) in comparison with Prorocentrum micans (GenBank access. no. X16108). The D2 region was the longest in length (300 bp) and highly variable among the 12 D-domains. In a phylogenetic analysis using complete LSU rDNA sequences of a variety of phytoplankton, A. tamarense was clearly separated with high resolution against other species. The result suggests that the sequence may resolve the taxonomic ambiguities of Alexandrium genus, particularly of the tamarensis complex.

• Journal of Microbiology
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• 제44권4호
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• pp.432-438
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• 2006

Twelve actinomycete strains were isolated from Egyptian soil. The isolated actinomycete strains were then screened with regard to their potential to generate antibiotics. The most potent of the producer strains was selected and identified. The cultural and physiological characteristics of the strain identified. the strain as a member of the genus Streptomyces. The nucleotide sequence of the 16S rRNA gene (1.5kb) of the most potent strain evidenced a 99% similarity with Streptomyces spp. and S. aureofaciens 16S rRNA genes, and the isolated strain was ultimately identified as Streptomyces sp. MAR01. The extraction of the fermentation broth of this strain resulted in the isolation of one major compound, which was active in vitro against gram-positive, gram-negative representatives and Candida albicans. The chemical structure of this bioactive compound was elucidated based on the spectroscopic data obtained from the application of MS, IR, UV, $^1H$ NMR, $^{13}C$ NMR, and elemental analysis techniques. Via comparison to the reference data in the relevant literature and in the database search, this antibiotic, which had a molecular formula of $C_{19}H_{29}NO_2$ and a molecular weight of 303.44, was determined to differ from those produced by this genus as well as the available known antibiotics. Therefore, this antibiotic was designated Meroparamycin.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2001년도 발생공학 국제심포지움 및 학술대회 발표자료집
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• pp.14-17
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• 2001

Positional cloning (map-based cloning) of mutations or genetic variations has been served as an invaluable tool to understand in-vivo functions of genes and to identify molecular components underlying phenotypes of interest. Mice homozygous for the cerebellar deficient folia (cdf) mutation are ataxic, with cerebellar hypoplasia and abnormal lobulation of the cerebellum. In the cdf mutant cerebellum approximately 40% of Purkinje cells are ectopically located within the white matter and the inner granule cell layer (IGL). To identify the cdf gene, a high-resolution genetic map for the cdf-gene-encompassing region was constructed using 1997 F2 mice generated from C3H/HeSnJ-cdf/cdf and CAST/Ei intercross. The cdf gene showed complete linkage disequilibrium with three tightly linked markers D6Mit208, D6Mit359, and D6Mit225. A contig using YAC, BAC, and P1 clones was constructed for the cdf critical region to identify the gene. A deletion in the cdf critical region on chromosome 6 that removes approximately 150kb of DNA was identified. A gene associated with this deletion was identified using cDNA selection. cdf mutant mice with the transgenic copy of the identified gene restored the brain abnormalities of the mutant mice. The positional cloning of cdf gene provides a good example showing the identification of a gene could lead to finding a new component of important molecular pathways.

• The Plant Pathology Journal
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• 제17권1호
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• pp.22-28
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• 2001

Barley yellow dwarf virus PAV (BYDV-PAV) has a 5.7-kb positive-sense single-stranded RNA genome that contains six open reading frames (ORFs). BYDV-PAV produces three subgenomic RNAs (sgRNAs). The largest of which encodes the coat, 17-kDa, and readthrough proteins from two initiation codons. To investigate the role of intergenic and 3'-proximal noncoding regions (NCRs) in coat protein (CP) expression and BYDV-PAV replication, a full-length infectious cDNA of the RNA genome of an Illinois isolate of BYDV-PAV was constructed downstream of the Cauliflower mosaic virus-35S promoter. Linear DNA molecules of these cDNAs were infectious, expressed the 22-kDa CP, and produced both genomic RNA sgRNAs in ratios similar to those observed in protoplasts inoculated with viral RNA. The portion of 5'NCR of sgRNA1 between ORFs 2 and 3 was not required for, but enhanced translation of CP from ORF3. Mutants containing deletions in the NCR downstream of ORF5 failed to replicate in oat protoplasts. These results indicate that an intact 3$^1$NCR is required for BYDV-PAV replication.

• 한국광물학회지
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• 제8권2호
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• pp.57-72
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• 1995

춘천 지역 선캠브리아기의 구봉산층군 내의 백운석질 대리암에는 광역변성작용의 산물로서 조립질의 투각섬석이 주된 규산염 광물상을 이루며 산출된다. 이들은 배태 층준과 원암의 조성에 따라 두 가지 유형으로 구분된다. : (1) 투각섬석(흑회색)+방해석+백운석+활석, (2)투각섬석(백색)+금운모+백운석+방해석+K-장석, 백색의 투각섬석이 흑회색형보다 상대적으로 높은 Al과 알칼리 함유도를 보이지만, 전반적으로 모두 Mg-단종에 가까운 화학 조성을 갖는다. 투각섬석들이 격자 구조상의 결함 구조를 갖지 않는데 비해서, 금운모는 1M 구조형을 이루고 저면 격자 간격상에 녹니석 단위의 빈번한 개재에 의한 격자상의 구조적 결함이 흔히 관찰된다.방해석-백운석 지온계에 대한 연구와 TWEEQU 프로그램을 이용한 상평형도 작성을 통해서, 투각섬석들은 500~$600^{\circ}C$의 온도 범위, 8kb 정도의 압력 및 Xco2=0.2~0.5 조건에서 생성된 것으로 해석하였다. 이 지역의 투각섬석은 위와 같은 조건에서 지속적으로 평형 상태가 유지되고 상안정도가 증대되므로서, (1) 투휘석을 수반하지 않고, (2) 격자 구조상의 결합을 갖지 않는, (3) 자형의 큰 결정을 이루게 된 것으로 해석된다.

• Journal of Microbiology and Biotechnology
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• 제14권1호
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• pp.197-201
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• 2004

A 2-D gel protein analysis of Lactobacillus reuteri ATCC 55739 produced spots corresponding to subunits of the pyruvate dehydrogenase complex, as identified by N-terminal protein sequencing. Oligonucleotide probes specific for the subunits of the pyruvate dehydrogenase complex were synthesized ,md used to screen a L. reuteri genomic library to clone the structural genes. Two positive clones were isolated and identified as having the same 2.2 kb insert. A pdhB encoding the $\beta$-subunit of El subunit (pyruvate dehydrogenase component) of the pyruvate dehydrogenase complex was located in the middle of the insert. Furthermore, a 5' truncated pdhA encoding the $\alpha$-subunit of the E1 subunit and a 3' truncated pdhC encoding the E2 subunit (dihydrolipoamide acetyltransferase) were also located upstream and downstream of the pdhB, respectively.

• 식물조직배양학회지
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• 제22권6호
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• pp.351-355
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• 1995

국내 재배종 토마토 서광 품종으로부터 분리한 Polygalacturonase 유전자(PG2)의 3'측 1.1 kb cDNA 단편을 식물 형질전환용 운반체에 antisense 방향으로 삽입한 후 자엽을 이용하여 토마토내 도입하여 형질전환 토마토를 획득하였다. 형질전환 토마토(T$^{0}$ )를 도입시켜 그 종자를 1 mg/mL 농도의 kanamycin 함유 MS 배지에서 발아시켜 분리 집단 중에서 T$_1$9 식물체를 얻었다. T$_1$9의 Genomic Southern blot 분석 결과, antisense PG 유전자 1개가 염색체 내로 삽입되었음을 확인하였고 RNA gel blot 분석으로 endogenous PG mRNA보다 antisense PG RNA가 강하게 발현됨을 확인하였다. T$_1$9 계통 10개체의 성숙 토마토 과피조직내의 PG 효소 활성도 4~60%까지 저해되었다.

• Journal of Plant Biotechnology
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• 제35권1호
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• pp.87-94
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• 2008

Porcine epidemic diarrhea virus (PEDV)는 돼지의 급성장염을 유발하여 설사 등의 증상을 일으키는 바이러스이다. 본 연구에서는 PEDV 항원단백질을 생산하는 담배 배양세포주를 개발하고자 하였다. PEDV에서 항원성이 알려진 스파크 단백질의 일부분을 암호화하는 유전자를 PCR로 합성하여 4종류의 형질전환 벡터를 제작하였다. 담배 배양세포 BY-2를 재료로 하여 Agrobacterium tumefaciens을 매개로 형질전환하였다. 선발배지 (MS salt, $KH_2PO_4$ 370 mg/L, 2,4-D 0.18 mg/L, Thiamin HCl 1 mg/L, kanamycin 100 mg/L, 침랙무 400 mg/L)에서 캘러스를 3주 간격으로 3개월 동안 계대배양하여 카나마이신 저항성 캘러스를 선발하였다. 선발된 캘러스를 대상으로 PCR 분석한 결과 형질전환 효율은 75% 이상이었으며 벡터당 40 여개 이상의 형질전환 배양세포주를 얻었다. 형질전환 배양세포주를 대상으로 Southern blot 분석하여 PEDV 유전자가 고구마 식물체의 게놈으로 안정적으로 도입되었음을 확인하였다. Northern blot 분석 결과 PEDV 스파크 단백질 유전자가 높은 수준으로 발현함을 확인하였으며 dot blot으로 PEDV 스파크 단백질 고생산 배양세포주를 선발하였다. 형질전환 담배 배양세포로부터 생산된 PEDV 항원단백질을 BALB/c 마우스에 경구투여 하여 면역활성을 조사한 결과 형질전환 세포주인 35S::SP1-M, 35S::SP2-M, 35S::SP4-M 세포주에서 1:10의 희석배수까지 바이러스 억제효과가 관찰되었다. 제작된 형질전환 벡터는 고구마와 같은 경구용 사료작물에 활용할 수 있을 것이다.