• 미생물학회지
• /
• 제32권1호
• /
• pp.20-27
• /
• 1994

항생물질에 대한 다중 저항성을 갖는 Staphylococcus aureus 균주의 chromosomal DNA로부터 유발성 발현을 하는 ${\beta}$-lactamase(bla) 유전자를 확인, 분리하였다. Cloning에 이어 결정된 염기서열을, S. aureus 에서는 지금까지는 plasmid상에서만 분리, 보고되어 있는 bla 유전자들의 염기서열과 비교하였다. 본 bla 유전자의 구조유전자 부분인 843base의 염기서열은 기 발표된 pPC1, pl258, pS1, pI1071, pUB101, pl3796 및 pI3804 유래의 bla 유전자들 중 pPC1, pI258 및 pS1상에 존재하는 bla 구조유전자의 염기서열과 완전히 일치하였고 나머지 것들과도 매우 높은 상동성(99%)을 유지하고 있었다. Bla구조 유전자의 상류 370base 및 하류 220base까지 결정된 염기서열을 비교한 결과에서는 다른 모든 bla구조유전자의 상류 150base에 위치하는 HindIII 인식부위가 약 230base 이상 더 윗쪽으로 옮겨가 있었고 이 HindIII 인식부위를 포함하는 염기서열에서 ORF의 C말단이 발견되었다. 하류 서열에서는 pI1071 유래 bla가 갖는 두개의 직접반복 염기서열 중 하나가 결손된 형태를 보이고 있다. 구조 유전자 상류에 존재하는, 80개 아미노산으로 구성된 ORF의 상동성 검색 결과 Tn4001 의 transposase 의 C 말단과 일치함이 발견되었다.

• The Plant Pathology Journal
• /
• 제20권3호
• /
• pp.229-233
• /
• 2004

The bacterial leaf blight, which is caused by Xantho-monas oryzae pv. oryzae, is the most damaging and intractable disease of rice. To identify the genes involved in the virulence mechanism of transposon TnS complex, which possesses a linearized transposon and transposase, was successfully introduced into X. oryzae pv. oryzae by electroporation. The transposon mutants were selected and confirm the presence of transposition in X. oryzae pv. oryzae by the PCR amplification of transposon fragments and the Southern hybridization using these mutants. Furthermore, transposon insertion sites in the mutant bacterial chromosome were deter-mined by direct genomic DNA sequencing using transposon-specific primers with ABI 3100 Genetic Analyzer. Efficiency of transposition was influenced mostly by the competence status of X. oryzae pv. oryzae cells and the conditions of electroporation. These results indicated that the insertion mutagenesis strategy could be applied to define function of uncharacterized genes in X. oryzae pv. oryzae.

• 생명과학회지
• /
• 제8권3호
• /
• pp.285-293
• /
• 1998

이미 밝혀져 있는 mariner 전이인자의 전이효소를 암호화하는 부위에 대하여 퇴화성 primer를 사용하여 PCR 방법에 의해 누에(Bombyx mori)에서 ariner 유사 전이닌자의 잠정적인 전이효소 부위를 클로닝 하였다. BmoMAR로 망명된 이 PCR 클론으로부터 추론된 아미노산은 152개로 다섯 개의 종결코돈이 삽입되어 있었으며, Drosophila mauritiana의 active Mos 1에 37%의 아미노산 상동성을 보였다. 또한, 기존의 곤충들에서 밝혀진 mariner-like element에 대한 상동성은 DNA 수주에서는 Apis mellifera에 59% 그리고 아미노산 수준에서는 D. mauritiana 7.9 clone에 37% 상동성을 보였다. 이 결과는 mariner-like element가 B. mori에도 존재하고 있지만. 이들 전이인자의 전이효소를 암호화하는 부위에 종결코돈이 발견되는 것으로 보아서 비활성 전이인자 혹은 일존의 selenoprotein으로 추정된다.

• BMB Reports
• /
• 제34권6호
• /
• pp.584-589
• /
• 2001

One of the cryptic plasmids from the oil degrading bacterium Klebsiella sp. KCL-2, the small plasmid pGD2, has been identified and characterized. This plasmid has a size of 3.6 kb with unknown functions. We constructed the recombinant plasmid pMGD2. The nucleotide sequences of the plasmid were determined and two open reading frames were detected. ORF1 encodes a replication initiator protein (RepA), which has a high degree of homology with the protein of ColE2 plasmid. The product encoded by ORF2 showed a high similarity with the transposase protein of IS5. IS5 is 1195 by long and contains an inverted terminal repetition of 16 bp with one mismatch. Stem-loop structures in the 5'untranslated region of the repA suggest that a putative gene, incA, is located in a complementary strand to the leader region of the repA mRNA.

• 한국식물병리학회:학술대회논문집
• /
• 한국식물병리학회 1995년도 Proceedings of special lectures on Molecular Biological Approaches to Plant Disease National Agricultural Science and Technology Institute Suwon, Korea
• /
• pp.1-19
• /
• 1995

The plasmid pJEL 101 contains a highly repetitive element from the genome of Xanthomonas oryae pv. oryzae that has properties of an insertional element. The insertional nature of the element, hereto referred to as IS203, was confirmed by molecular analyses of the element and three related elements that were isolated from X. oryzae. The related sequences were isolated on the basis of transposition to the transposon-trapping vector pL3SAC and hybridization with pJEL101. The trapped elements (IS203a, IS203b, and IS203c) were each composed of 1,055 base pairs with 25 base terminal inverted repeats. The elements caused a three base pair target site duplication at the site of insertion in the sacRB gene. The sequence of pJEL 101 has 96% base pair identity with IS203a and 99% identity with IS203a and IS203c but lacks three nucleotides of the consensus left terminal repeat. IS203b has the same DNA sequences as IS203c but is inserted ito the sacRB gene in the opposite orientation. The longest open reading frame of IS203a could code for a protein of 318 amino acids and molecular weight of 37, 151. A search of the Genbank database revealed that IS203 has 51% identity with 909 nucleotides of IS4551 from Escherichia coli. The predicted protein of ORF1 has 40% and 30% amino acid identity to the ORF1 of Tn4551 and the transposase of IS30, respectively.

• The Plant Pathology Journal
• /
• 제32권6호
• /
• pp.575-579
• /
• 2016

Burkholderia glumae (bacterial grain rot), Xanthomonas oryzae pv. oryzae (bacterial leaf blight), and Acidovorax avenae subsp. avenae (bacterial brown stripe) are major seedborne pathogens of rice. Based on the 16S and 23S rDNA sequences for A. avenae subsp. avenae and B. glumae, and transposase A gene sequence for X. oryzae pv. oryzae, three sets of primers had been designed to produce 402 bp for B. glumae, 490 bp for X. oryzae, and 290 bp for A. avenae subsp. avenae with the $63^{\circ}C$ as an optimum annealing temperature. Samples collected from naturally infected fields were detected with two bacteria, B. glumae and A. avenae subsp. avenae but X. oryzae pv. oryzae was not detected. This assay can be used to identify pathogens directly from infected seeds, and will be an effective tool for the identification of the three pathogens in rice plants.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제9권2호
• /
• pp.183-186
• /
• 2004

Recently, we have developed an easy, simple and convenient circular DNA cloning system named plasmid capture system (PCS). To investigate usefulness of PCS in cloning of plasmids from Bacillus thuringiensis strains, PCS donors, pPCS-S and pPCS-L were applied to clone plasmids of B. thuringiensis subsp. israelensis by in vitro transposition using 4{TnsABC^*}$ transposase. In result, 3 small plasmids were cloned, and these were consistent with pTX14-1, pTX14-2 and pTX14-3 reported previously from B. thuringiensis subsp. israelensis. Therefore, the PCS can be successfully applied to clone small plasmids from B. thuringiensis strains.

• Asian-Australasian Journal of Animal Sciences
• /
• 제30권6호
• /
• pp.886-892
• /
• 2017

Objective: Transgenic technology is widely used for industrial applications and basic research. Systems that allow for genetic modification play a crucial role in biotechnology for a number of purposes, including the functional analysis of specific genes and the production of exogenous proteins. In this study, we examined and verified the cumate-inducible transgene expression system in chicken DF1 and quail QM7 cells, as well as loxP element-mediated transgene recombination using Cre recombinase in DF1 cells. Methods: After stable transfer of the transgene with piggyBac transposon and transposase, transgene expression was induced by an appropriate concentration of cumate. Additionally, we showed that the transgene can be replaced with additional transgenes by co-transfection with the Cre recombinase expression vector. Results: In the cumate-GFP DF1 and QM7 cells, green fluorescent protein (GFP) expression was repressed in the off state in the absence of cumate, and the GFP transgene expression was successfully induced in the presence of cumate. In the cumate-MyoD DF1 cells, MyoD transgene expression was induced by cumate, and the genes controlled by MyoD were upregulated according to the number of days in culture. Additionally, for the translocation experiments, a stable enhanced green fluorescent protein (eGFP)-expressing DF1 cell line transfected with the loxP66-eGFP-loxP71 vector was established, and DsRed-positive and eGFP-negative cells were observed after 14 days of co-transfection with the DsRed transgene and Cre recombinase indicating that the eGFP transgene was excised, and the DsRed transgene was replaced by Cre recombination. Conclusion: Transgene induction or replacement cassette systems in avian cells can be applied in functional genomics studies of specific genes and adapted further for efficient generation of transgenic poultry to modulate target gene expression.

• 생명과학회지
• /
• 제9권2호
• /
• pp.176-181
• /
• 1999

본 연구는 누에(Bombyx mori)에서 cloning한 BomMAR의 염기서열을 기초로 하여 곤충 MLE(mariner-like ele-ment)의 계통분석을 통한 누에의 분자적 계통 관계를 이해하고자 수행하였다. 전체 10 종의 MLE중에서 15%의 낮은 상동성을 보이는 인간 MLE(Hsmarl)을 제외한 9종의 MLE의 DNA 염기서열을 이용한 UPGMA 분석 결과, BmoMAR을 포함한 10종의 MLE가 세 가지의 subfamily 로 grouping되는 것을 알 수 있었으며, 누에는 genetic distance 0.4332에서 같은 lepidoptera(나비목) 의 H. cecropia, almond moth, webworm 및 microcaddisfly와 함께 grouping 되었다. 따라서 이와 같은 결과는 MLE가 곤충의 계통분석에 유용한 molecular tool이 될 수 있음을 보여준다.

• The Plant Pathology Journal
• /
• 제36권5호
• /
• pp.428-439
• /
• 2020

Erwinia pyrifoliae is a Gram-negative bacterial plant pathogen that causes black shoot blight in apple and pear. Although earlier studies reported the genome comparison of Erwinia species, E. pyrifoliae strains for such analysis were isolated in 1996. In 2014, the strain E. pyrifoliae EpK1/15 was newly isolated in the apple tree showing black shoot blight in South Korea. This study aimed to better understand the similarities and differences caused by natural variations at the genomic level between newly isolated E. pyrifoliae EpK1/15 and the strain Ep1/96, which were isolated almost 20 years apart. Several comparative genomic analyses were conducted, and Clusters of Orthologous Groups of proteins (COG) database was used to classify functional annotation for each strain. E. pyrifoliae EpK1/15 had similarities with the Ep1/96 strain in stress-related genes, Tn3 transposase of insertion sequences, type III secretion systems, and small RNAs. The most remarkable difference to emerge from this comparison was that although the draft genome of E. pyrifoliae EpK1/15 was almost conserved, Epk1/15 strain had at least three sorts of structural variations in functional annotation according to COG database; chromosome inversion, translocation, and duplication. These results indicate that E. pyrifoliae species has gone natural variations within almost 20 years at the genomic level, and we can trace their similarities and differences with comparative genomic analysis.