• Current Research on Agriculture and Life Sciences
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• 제5권
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• pp.52-58
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• 1987

효모(酵母)와 대장균(大腸菌)의 셔틀 벡터인 YIp5, YRp7, YEp13플라스미드를 S. cerevisiae DBY747을 수용세포(受容細胞)로 하여 원형질체 방법(方法)에 의해 형질전환(形質轉換)시킨 결과(結果), YIp5플라스미드는 형질전환체가 나타나지 않았으며, YEp13플라스미드는 DNA $10{\mu}g$ 당(當) $1.2{\times}10^3$, YRp7은 $1.0{\times}10^2$ 개(個)의 형질전환체를 얻었다. YIp5플라스미드와 YEp13플라스미드, 그리고 YIp5플라스미드와 YRp7플라스미드를 환상(環狀)플라스미드 상태(狀態)로 동시형질전환(同時形質轉換) 시켰을 때 각각 DNA $10{\mu}g$ 당(當) 210 및 95개(個)의 형질전환체를 얻었으며, 동일(同一) 부위(部位) 또는 다른 부위(部位)를 절단(切斷)한 선상(線狀)플라스크를 앞서와 같이 동시형질전환(同時形質轉換)시켰을때, 서로 비슷하게 DNA $10{\mu}g$당(當) 15~85개(個)의 형질전환체를 얻을 수 있었다. 이러한 결과(結果)에서 볼때 수용세포(受容細胞)를 S. cerevisiae DBY747로한 동시형질전환(同時形質轉換)에서 환상(環狀)플라스미드간(間)의 형질전환(形質轉換)이 선상(線狀)플라스미드간(間)의 형질전환(形質轉換)보다 빈도(頻度)가 높으며 선상(線狀)플라스미드간(間)의 형질전환(形質轉換)에서 미단(未端) 부위(部位)의 상이(相異)함이 큰 영향을 주지 않는 것으로 나타났다. 형질전환체의 안정성(安定性)에 있어서는 YIp5플라스미드와 YEp13플라스미드간(間)의 형질전환체가 YIp5플라스미드와 YRp7플라스미드간(間) 형질전환체에 비(比)해 안정(安定)하게 나타났는데, 이는 stringent control을 받는 ARS유전자(遺傳子)를 가진 YRp7플라스미드보다 $2{\mu}m$ DNA 복제(複製) 기점(起點)을 가진 YEp13플라스미드의 복제수(copy number)가 많기 때문인 것으로 생각된다.

• Journal of Microbiology and Biotechnology
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• 제4권1호
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• pp.1-6
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• 1994

Continuous culture was carried out with a recombinant Escherichia coli W3110/pCR185, which encodes trp-operon enzymes when the temperature is shifted from $37^{circ}C\;t;42^{\circ}C$. Under derepressed condition of $42^{\circ}C$. plasmlid stability and gene expression were analysed as function of the dilution rate. The stability of plasmid increased with the dilution rate, but maximal levels of gene expression (tryptophan concentration) and plasmid DNA content were obtained at the lowest dilution rate, $0.075\;hr^{-1}$. The plasmid instability, observed at low dilution rates, could be explained by the unbalanced biosynthetic state of the recombinant cell harboring a high copy number of plasmid.

• 한국미생물·생명공학회지
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• 제20권3호
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• pp.280-288
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• 1992

Serratia marcescens ATCC 21074 균주가 세포밖으로 분비하는 metalloprotease 유전자를 대장균으로 클로닝하고 그 발현을 살펴보았다 Serratia marcescens ATCC 21074 균주의 염색체 DNA를 제한효소 HindIII로 절단하고 아가로스 전기영동 후 32P로 표지된 합성 oligonucleotide를 사용하여 southern hybridization한 결과 4.0Kb의 DNA 절편에 metalloprotease가 존재함을 알 수 있었다. 4.0Kb 염색체 DNA 절ㅊ편을 분리하여 pUC19에 연결한 후 대장균으로 transformation하였다.

• Fisheries and Aquatic Sciences
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• 제7권2호
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• pp.70-75
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• 2004

The CMV promoter driven lacZ reporter gene (pcDNA-lacZ) was constructed and used for DNA immunization study. The expression of the lacZ gene was confirmed in vitro using RTG-2 cell line before using for in vivo study in olive flounder (Paralichthys olivaceus). In the dose response study, the maximum expression of the lacZ gene was found in the group injected with 5 ${\mu} g$ of the plasmid DNA. Kinetic study showed a significantly increased expression of $\beta-galactosidase$ gene at 7 days after injection. Effects of DNA vaccine on specific and nonspecific immune responses such as antibody and NO production were studied and the significant effect was found in olive flounder injected with 10 and 15 ${\mu} g$ DNA (sub optimal dose for lacZ gene expression). Two pro inflammatory cytokine genes, $IL-l\beta$ and $TNF-\alpha$, were also found to be up regulated in the muscle injected with the plasmid, suggesting an induction of local inflammatory response.

• 한국가시화정보학회:학술대회논문집
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• 한국가시화정보학회 2003년도 추계학술대회 논문집
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• pp.81-82
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• 2003

We have developed the method for the conjugation of biotinylated DNA to streptavidin-coated QDs. QD-DNA conjugates and a high-sensitive fluorescence imaging technique are adopted to visualize gene transport across the membrane of the live cell in real time. Endocytotic cellular uptake of oligonucleotide and electrically-mediated plasmid DNA transfer into the live cell are monitored by a quantitative microscopic imaging system. Long-term kinetic study enables us to reveal the unknown mechanisms and rate-limiting steps of extracellular and intracellular transport of biomolecules. We designed experimental protocols to conjugate the oligonucleotide or the plasmid DNA to commercially available streptavidin-coated QDs. Gel electrophoresis is used to verify the effect of incubation time and the molar ratio of QDs and DNA on the conjugation efficiency. It is possible to fractionate the QD-DNA conjugates according to the DNA concentration and obtain the purified conjugates by a gel extraction technique.

• 한국연초학회지
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• 제19권1호
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• pp.11-17
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• 1997

A flue-cured tobacco variety (Nicotiana tabacum cv. Wisconsin) was used for Plant transformation with the complementary DNA (cDNA) of potato virus Y-necrosis strain (PVY-VN) replicase gone (Nb) which was synthesized through reverse-transcription Primed with oligo(dT) and Polymerization using RNase H-digested template. The cDNA was cloned into Plant expression vector Plasmid (PMBP2), and introduced into tobacco plants by co-culturing tobacco leaf disks with Agrobacterium tumefaciens LBA4404 containing the plasmid before Plant regeneration. Eight Plants, in which the inserted cDNA fragment was detected by Polymerase chain reaction (PCR), out of 70 putative transformants inserted with sense-oriented Mb cDNA showed no symptom at 3 weeks after inoculation, while the other 62 plants, and all plants with vector gone only and antisense-oriented NIb cDNA had susceptible vein-necrosis symptoms. However, only 2 of the 8 resistant plants were highly resistant, which remained symptomless up to 10 weeks after inoculation. Among the first progenies (T1) from self-fertilized seeds of the two resistant transgenic plants, less than 10 % of 71 plants appeared highly resistant (with no symptom), 70% moderately resistant (with mild symptoms on 1 - 2 leaves), and about 20% susceptible (with susceptible symptoms on 3 or more leaves) at 3 weeks after inoculation. These results suggest that the PVY resistance was inherited in the 71 generation. Key words : potato virus Y. viral replicase gene, transgenic tobacco Plants, resistance.

• 한국미생물·생명공학회지
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• 제14권6호
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• pp.455-460
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• 1986

간염 보균자의 혈액으로부터 Dane 입자를 분리하였다. Dane 입자의 핵으로부터 분리해낸 DNA는 $\alpha$-($^{32}$P) dNTP 존재하의 DNA 폴리머레이즈 반응 후 액체 씬틸레이션 카운터와 한천 전기영동 및 가이거 뮐러 카운터에 의하여 간염의 DNA임이 확인되었다. 간염 바이러스에 의한 감염을 막기 위한 백신으로서의 B형 간염 바이러스 표면항원을 생산하기 위하여 산성포스파테이즈 프로모터를 갖는 재조합 프라스미드를 함유하는 효모균주를 사용하였다. 재조합 프라스미드는 pHBV 130 및 pAM 82로부터 제작되었으며 대장균에 변환되어진 후 효모균주에 전달되었다. 간염 표면항원은 조절된 무기 인산 농도하에서 버크홀더 최소배지에서의 저해 해제로 생산되었다. 간염 표면항원의 생산 속도도 조사하였다. 전체 간염 표면항원 활성은 인산이 없는 배지에 옮겨진 뒤 3시간 내지 6시간에서 급격히 증가하였으며 9시간째에 최대에 도달하였다. 인산이 없는 배지에 옮기는 것은 고농도 인산 배지에서의 세포 배양을 6시간동안 수행한 뒤에 하는 것이 최적의 결과를 나타내었다.

• 미생물학회지
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• 제24권1호
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• pp.12-17
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• 1986

A new computer algorithm is described to order the restriction fragments of plasmid DNA which has been cleaved with several restriction endonucleases in single or double digestions rapidly with realistic error rates. The permutation and high weight on small fragments methods construct all logical circular map solutions. The program is written in Apple BASIC and run on an Apple II plus microcomputer with 64K memory. Several examples are presented which indicate the high efficiency of the profram in construction possible restriction map for YEp24.

• 한국축산식품학회지
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• 제38권4호
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• pp.816-822
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• 2018

Bifidobacterium is recognized as one of the most beneficial microorganisms in our gut. Many researches on bifidobacteria have been done to understand their roles in the gut. The objective of the present study was to develop a bioluminescent labelling plasmid vector for bifidobacteria to facilitate their visualization in vitro, in situ, and in vivo. A plasmid replicon (2.0 kb) of plasmid pFI2576 previously identified from B. longum FI10564 was amplified by PCR and cloned into pUC19 plasmid vector (2.68 kb). The cloned replicon was subcloned into pTG262 ($luc^+$) recombinant plasmid vector (7.4 kb) where a luciferase gene ($luc^+$) from pLuc2 (8.5 kb), an Escherichia coli and lactobacilli shuttle vector, was inserted into pTG262 plasmid vector. The final recombinant DNA, pTG262::pFI2576 rep ($luc^+$), was transferred into a B. catenulatum strain. This recombinant strain showed 3,024 relative luminescence units at $OD_{600}$ value of 0.352. Thus, this recombinant plasmid construct can be broadly used for labelling bifidobacteria.

• 한국어병학회지
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• 제12권2호
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• pp.115-121
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• 1999

양식넙치에서 분리된 Edwardsiella tarda가 갖는 약제내성을 전달하는 R plasmid의 특성을 알아보기 위하여, 16종의 화학요법제에 대한 E. tarda의 감수성 정도를 비교하였으며, 내성형태 및 내성전달을 확인하고 transferable R plasmid를 분리하였다. E. tarda는 ampicillin, amoxicillin, erythromycin, flumequine, doxycycline(DOXY), nalidixic acid, novobiocin, oxolinic acid, oxytetracycline(OTC), thiamphenicol(TP) 그리고 sulfonamide의 11약제에 대하여 다양한 조합으로 복합내성을 보였으며, DOXY, OTC, TP 내성이 Escherichia coli로 전달되었다. 복합내성의 두 균주에서 transconjugant가 형성되었으며 이들로부터 분리된 transferable R plasmid는 서로 상이한 DNA 구조였다. 이는 넙치에서 분리되는 E. tarda에는 적어도 두 종류 이상의 thansferable R plasmid가 분포한다는 것을 의미한다.