• Journal of the korean society of oceanography
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• 제39권3호
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• pp.163-171
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• 2004

Sequences of the large subunit ribosomal (LSD) rDNA D1-D2 region of Alexandrium catenella(=A. sp. cf. catenella) and A. tamarense isolates, which were collected along the Korea coasts, were analyzed to understand their phylogenetic relationships and geographical distributions. All A. catenella and A. tamarense isolates belonged to the A. tamarense/catenella/fundyense complex and were grouped with the North American and temperate Asian ribotypes, respectively, regardless of the presence or absence of a ventral pore in the first apical plate. A consistent and peculiar characteristic that differentiated the Alexandrium isolates was amplification of a second PCR product with a lower molecular weight in addition to the predicted one; ten A. catenella isolates belonging to the temperate Asian ribotype yielded this additional PCR product. Sequence alignment revealed that the shorter PCR product resulted from an unusual large deletion of 87 bp in the LSD rDNA D1 domain. The North American and temperate Asian ribotypes were prevalent along the Korean coasts without geographical separation. Given the high genetic homogeneity among widely distributed Alexandrium populations, each ribotype appeared to be pandemic rather than to constitute a distinct regional population.

• 한국식품과학회지
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• 제33권5호
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• pp.521-524
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• 2001

본 연구는 유전자재조합 기술에 의해 개발된 glyphosate에 내성을 가지고 있는 콩(GTS)의 모니터링을 위하여 PCR을 이용한 검출 방법에 대한 실험을 수행하였다. Glyphosate에 내성이 있는 콩에 삽입된 유전자와 표준대조 유전자인 lectin과 ferritin 유전자를 근거로 제작된 primer와 CTAB 방법으로 추출된 콩의 DNA를 template로 이용하여 PCR을 수행하였다. GTS의 검출을 위한 제작된 primer들은 GTS와 특이적으로 반응하여 증폭된 PCR 산물을 생성하였으나, non-GTS와는 PCR 산물을 생성하지 못했다. 증폭된 염기서열 분석을 통하여 GTS에 특이적인 것을확인하였으며, 약 0.05%가 포함되어 있는 GTS까지 검출이 가능함을 보였다.

• 생명과학회지
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• 제9권1호
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• pp.15-21
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• 1999

PCR 기법을 응용한 RAPD 방법은 대상 생물의 유전정보를 전혀 모르는 상태에도 arbitrary primer를 이용하여 증폭함으로써 생물 종간의 유전적 표식자를 확인할 수 있는 간편하고 유익한 유전분석 방법이다. 이같은 RAPD 방법을 이용하여 해조류 방사무늬 김, 미역, 구멍갈파래에 대하여 DNA를 추출하지 않고 직접 생 엽체 및 생 사상체, 생 배우체를 PCR template로 사용하여 PCR product를 생산하였다. 그러나 specific primer를 이용한 nulear r DNA의 internal trancribed spacer (ITS) 부위는 생성물을 만들지 못하였다. 간편한 LiCl 방법에 의하여 추출된 DNA를 사용하였을 때는 IST 및 RAPD 모두 PCR product를 생산하였다. 방사무늬 김의 엽체 (haploid)와 사상체 (dip-loid)로 부터의 ITS 생성물은 동일하였으나 RAPD 생성물은 사용한 arbitrary primer에 따라 36-50$\%$의 다른 band가 만들어졌다. 또한 직접 생 조직을 사용하였을때와 추출 DNA를 사용하였을 때도 53-57$\%$의 다른 band가 만들어 줬다. 그러므로 RAPD 방법으로 유전분석 실험에는 동일한 ploidy의 조직을 template로서 사용하는 것이 중요하다.

• 한국미생물·생명공학회지
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• 제32권3호
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• pp.282-285
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• 2004

The polyene antibiotics including nystatin, pimaricin, amphotericin and candicidin are a family of most promising antifungal polyketide compounds, typically produced by rare actinomycetes species. The biosynthetic gene clusters for these polyenes have been previously investigated, revealing the presence of highly homologous biosynthetic genes among polyene-producers such as polyketide synthase (PKS) and cytochrome P450 hydroxylase (CYP) genes. Based on amino acid sequence alignment among actinomycetes CYP genes, the highly-conserved regions specific for only polyene CYP genes were identified and chosen for degenerate PCR primers, followed by the PCR-screening with various actinomycetes genomic DNAs. Among tested several polyene non-producing actinomycetes strains, Pseudonorcardia autotrophica strain was selected based on the presence of PCR product with polyene-specific CYP gene primers, and then confirmed to contain a cryptic novel polyene hydroxylase gene in the chromosome. These results suggest that the polyene-specific hydroxylase gene PCR should be an efficient way of screening and isolating potentially-valuable cryptic polyene antibiotic biosynthetic genes from various microorganisms including rare actinomycetes.

• The Plant Pathology Journal
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• 제16권1호
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• pp.48-51
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• 2000

Citrus tristeza virus (CTV) was identified form CTV-infected early satsuma mandarin (Citus unshiu) and yuzu (C.junos) by RT-PCR. The total RNAs were isolated from citrus bark and seaf tissues infected with CTV and reverse transcription was followed with primers designed for amplifying CTV coat protein gene. DNA fragments 738 bp were amplified by RT-PCR and these products were colned for sequence analysis. Based on the sequence analysis, this PCR product has 97% sequence homology to CTV (T-385) CP gene isolated from USA. RT-PCR assay for CTV detection was more sensitivity than ELISA assay which was done with anti-CTV CP antibody. This is the frist report about CTV identification in Cheju island Korea.

• Mycobiology
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• 제31권1호
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• pp.19-22
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• 2003

The primary objective of the current research was to detect genetic variations within the Indonesian isolates of Cochliobolus heterostrophus collected from ecologically different places of the country at molecular level using PCR-RFLP analyses. The primer pair of NS3 and NS6 produced amplification fragment in all of the isolates tested. A single fragment of estimated 907 bp was observed in the PCR product pattern. RFLP analysis of the PCR product employing three restriction enzymes, HaeIII, HhaI, and RsaI, respectively, did not reveal intraspecific variations within the fungus. Similarly, nucleotide sequences of portion of small subunit of the ribosomal DNA gene of two of the isolates collected showed no appreciable differences, indicating the absence of genetic diversities among the isolates tested. A phylogenetic tree was constructed and the Indonesian C. heterostrophus, represented by SM-1 isolate, was found to be phylogenetically located near C. sativus, a closely related species.

• BMB Reports
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• 제29권1호
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• pp.52-57
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• 1996

A specific DNA fragment from Korean radish (Raphanus sativus L.) was amplified by performing PCR with oligonucleotide primers which correspond to the highly conserved regions of plant peroxidases. The size of the PCR product was ca. 400 bp, as expected from the known plant peroxidase genes. Comparison of the nucleotide and deduced amino acid sequences of the PCR product to those of other plant peroxidase-encoding genes revealed that the amplified fragment corresponded to the highly conserved region I and III of plant peroxidases. By screening a genomic library of Korean radish using the amplified fragment as a probe, two positive clones, named prxK1 and prxK2, were isolated. Restriction mapping studies indicated that the 5.2 kb Sail fragment of the prxK1 clone and the 4.0 kb EcoRI fragment of the prxK2 clone encode separate isoperoxidase genes. Analyses of the promoter region of the prxK1 clone shows that putative CAAT box, CMT box, and TGA1b binding sequence (5' TGACGT) are present 718 bp upstream from the start codon.

• 한국약용작물학회지
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• 제18권1호
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• pp.40-45
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• 2010

The application of PCR analysis on the herbal medicine Moutan Cortex (Paeonia suffruticosa Andrews) was evaluated by the comparison of the genetic relationship based on the DNA sequence with Paeoniae Radix (Paeonia lactiflora Pallas) following development of specific primers. Moutan Cortex and Paeoniae Radix were distinguished through the PCR analysis based on the internal transcribed spacer (ITS-PCR) from nuclear ribosomal DNA region. The 294 bp PCR products both of Moutan Cortex and Paeoniae Radix was amplified by MIF1 and MIR1. And a Moutan Cortex specific 225 bp PCR amplification product was amplified by MIF2 and MIR1 primers. The 225 bp sequence could be successfully amplified from Mortan Cortex of dried herbal preparations. PCR analysis based on ITS (ITS-PCR) may be an efficient tool for the discrimination of Moutan Cortex.

• Applied Biological Chemistry
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• 제39권2호
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• pp.112-117
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• 1996

ADC는 diamine인 putrescine 생합성의 두가지 경로중에서 식물계에서 특히 중요한 효소이며, ADC 유전자는 E. coli, 귀리, 토마토 genome에서 이미 cloning된 바 있다. 벼 (Oryza sativa L.) 게놈 DNA의 PCR 증폭을 위해서 토마토와 E. coli의 ABC cDNA의 보존된 부분과 일치하는 두개의 degenerate oligonucleotides (17mer)를 인위 합성하였으며, 증폭의 결과 약 1 kbp 크기의 DNA가 관찰되었다. 증폭된 DNA 절편은 1,022bp 염기서열을 포함하고 있는 ORE (open reading frame)으로 확인되었다. 이 PCR product는 POEM-originated T vector에 재조합하였으며 PstI 제한효소로 약 500bp 크기로 절단하여 pGEM-3Zf(+/-) vector에 subcloning하였다. 벼 ADC clone의 염기서열은 귀리와 토마토 ADC cDNA 서열의 같은 부분과 각각 74%와 70%의 동질성을 갖는 것으로 나타났으며, 예상되는 아미노산 서열은 귀리와 토마토 ADC 단백질과 각각 45%와 62%의 동질성이 관찰되었다. 귀리와 E. coli, 토마토와 귀리 그리고 토마토와 E. coli ADC 아미노산 서열에서 각각 34%, 47%, 그리고 38%의 유사성 정도가 보고된 것을 비교하여 볼 때, 벼와 귀리 및 토마토 사이의 유사성 정도는 다른 비교 보다도 월등히 높았다. 벼 유묘기 잎조직에서 추출한 RNA를 이용한 Northern blot 분석에서 ADC는 약 2.5kbp의 전사체로 발현됨이 확인되었다.

• 식물병연구
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• 제10권4호
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• pp.310-312
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• 2004

Pseudomonas syringae pv. actinidiae는 참다래 궤양병을 일으키는 세균이다. 식물독소인 coronatine 생합성에 관여하는 유전자 중 하나인 cfl의 염기서열로부터 설계된 primer를 사용한 nested PCR 방법을 토양시료에 적용시켰다. 이 primer 세트와 우리나라에서 분리된 P. syringae pv. actinidiae가 접종된 토양으로부터 얻은 DNA로 두 번의 PCR을 행했을 때 665 bp와 310 bp의 절편이 각각 증폭되었다. 이 시스템을 참다래 궤양병으로 폐원된 과수원의 토양조사에 적용시킨 결과 여섯 곳으로부터 채취한 토양시료 모두에서 특이적인 310 bp의 PCR 산물이 증폭되었다.