• 제목/요약/키워드: PCR-Hybridization

검색결과 437건 처리시간 0.029초

내열성 유전자 DgP23을 도입한 형질전환 오차드그라스의 생산 (Production of Transgenic Orchardgrass Overexpressing a Thermotolerant Gene, DgP23)

  • 김기용;장요순;박근제;최기준;성병렬;서성;차준영;손대영
    • 한국초지조사료학회지
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    • 제25권4호
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    • pp.267-274
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    • 2005
  • 고온 내성 오차드그라스를 개발하기 위하여, 재조합 DgP23 유전자에 CaMV 35S 프로모터를 붙여 발현벡터를 제작, Agrobacterium 형질전환 방법으로 오차드그라스 형질전환체를 생산하였다. Genomic DNA를 분리하여 PCR 및 Southern blot 분석을 실시한 결과, PCR 분석에서 DgP23 유전자의 DNA band가 관찰되었고, Southern blot 분석에서도 X-ray film 상에 hybridization signal이 관찰되어, 오차드그라스 genome에 DgP23 유전자의 도입이 확인되었으며, wild type 및 empty vector control에서는 DNA band 및 hybridization signal이 관찰되지 않았다. 또한 RT-PCR 및 이들 산물의 Southern blot 분석 결과, DgP23 유전자의 정상적인 발현이 확인되었다. 형질전환 오차드그라스를 온실 및 포장에서 재배하며 생육특성을 조사한 결과, 비형질전환체와 비교하여 형태적 차이는 나타내지 않았다. 실험실 조건에서 고온내성을 조사한 결과, 고온내성이 확인되지 않았기 때문에 형질전환 종자를 생산하여 포장조건에서 고온내성을 검정할 계획이며, 재배시험에서는 내성이 강한 개체를 선발할 수 있을 것으로 예상된다.

Development of Geno-chromatographic Assay System for Salmonella species

  • 김석하;오홍일;조정환;백세환
    • 한국생물공학회:학술대회논문집
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    • 한국생물공학회 2001년도 추계학술발표대회
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    • pp.827-828
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    • 2001
  • 이 실험은 특정 DNA를 검출 할 수 있는 새로운 방법을 제시하고 개발하려는 것이다. 이것은 geno-chromatography 분석방법이라고 하여 DNA capture probe를 membrane상에 고정하고 PCR을 통하여 증폭된 특정 유전자를 hybridization 반응을 이용하여 탐지하는 것이다. 이러한 개념을 식중독균의 일종인 Salmonella 균의 탐지에 적용하여 그 분석시스템의 성능을 확인 할 수 있었다.

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Follow-Up of Exogenous DNA by Sperm-Mediated Gene Transfer Via Liposome

  • Jo Hwang-Yun;Jo Seong-Geun;Yun Hui-Jun;Park Mi-Ryeong;Im Yeo-Jeong;Park Jong-Ju;Kim Jin-Hoe
    • 한국동물번식학회:학술대회논문집
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    • 한국동물번식학회 2002년도 춘계학술발표대회 발표논문초록집
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    • pp.86-86
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    • 2002
  • To examine the feasibility of using a sperm vector system for gene transfer, we have investigated the binding and the uptaking of foreign DNA into the sperm nucleus by PCR, in situ hybridization and LSC. We have also examined the transportation of exogenous DNA into oocytes by immunofluorescene via PCR. Sperm cells were incubated with DNA/liposome complexes (1:4 ratio) in fertilization medium with BSA or without BSA. (omitted)

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Effects of 17 β -estradiol, bisphenol A and genistein on the expression of the glutathione peroxidase gene of Philasterides dicentrarchii (Ciliophora: Scuticociliata)

  • 이은혜;김성미;남윤권;김기홍
    • 한국어병학회지
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    • 제19권3호
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    • pp.189-195
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    • 2006
  • A subtracted cDNA library of a marine scuticociliate, Philasterides dicentrarchii, in response to 17β-estradiol exposure was constructed using suppression subtractive hybridization (SSH). As a result of SSH, 275 clones were isolated, and among them, only glutathione peroxidase (GPX) gene was isolated as an antioxidative enzyme responding to 17β-estradiol. The semi-quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) analysis revealed that the transcription of GPX gene of P. dicentrarchii was clearly increased by exposure to 17β-estradiol. The GPX transcription was also clearly increased by exposure to xenoestrogens such as bisphenol A (BPA) and genistein.

Development of Nested RT-PCR for the Detection of Swine Hepatitis E virus in Formalin-fixed, Paraffin-embedded Tissues and Comparison with in situ Hybridization

  • Park, Jung-eun;Ha, Seung-kwon;Chae, Chan-hee
    • 한국수의병리학회:학술대회논문집
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    • 한국수의병리학회 2003년도 추계학술대회초록집
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    • pp.26-26
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    • 2003
  • Hepatitis E virus (HEV) has been recognized as a major cause of enterically transmitted non-A, non-B hepatitis in many developing countries. The taxonomy of HEV is not clear and the virus remains unclassified. The objective of this study was to optimize conditions and procedures to detect swine HEV in formalin-fixed, paraffin-embedded tissues by nested RT-PCR and compare this detection method with in situ hybridization. (omitted)

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Transovarial Transmission of Bombyx mori Nuclear Polyhedrosis Virus in the Silkworm

  • Xiao, Qing-Li;Zhang, Zhi-Fang;Yi, Yong-Zhu;He, Jia-Lu
    • International Journal of Industrial Entomology and Biomaterials
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    • 제2권2호
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    • pp.119-122
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    • 2001
  • Whether Bombyx mori nuclear polyhedrosis virus (BmNPV) can be transmitted to offspring, has been a noticeable question for a long time. When fifth instar larvae of the silkworm were orally inoculated with BmNPV dot hybridization and PCR amplification analysis demonstrated that BmNPV was not detected in the eggs laid by BmNPV productively infected female moths. The results indicated that BmNPV could not be transovarially transmitted.

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Genetic Distance of Allium Section Cepa by DNA Fingerprint

  • Kim, Haeng-Hoon;Cho, Eun-Gi;Baek, Hyung-Jin;Kim, Chang-Yung;Chae, Young-Am
    • 한국작물학회지
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    • 제48권1호
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    • pp.31-37
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    • 2003
  • Identification of compatible parental line is of great importance in introduction of useful characters to onion breeding program, beyond the severe hybridization barrier. Phylogenic analysis of Allium section Cepa was conducted through PCR by URPs, repeated sequences of A. fistulosum, and microsatellite markers. Totally 76 accessions originated from 21 countries were clustered into five groups at a 0.84-similarity level: group I;A. cepa and its wild relatives and A. cepa ssp. ascalonicum, group II; A. cepa ssp. wakegii, A. cepa ssp. proliferum and Samcheung-pa group III; A. fistulosum and A. altaicum, group IV; A. galanthum, group V; Soeckkori-pa. Samcheung-pa and Soekkori-pa, Korean local varieties, shared band type of both Cepa group and Altaicum group, indicating that those are derived from interspecific hybridization between A. fistulosum and A. cepa.

Evaluation of Several Parameters of in situ Polymerase Chain Reaction (ISPCR) to Reduce the Leakage of Amplificants from Cells

  • Lee, Jae-Yung;Auh, Chung-Kyoon;George W. Jordan
    • Journal of Microbiology
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    • 제40권1호
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    • pp.70-76
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    • 2002
  • Proviral DNAs from HIV-1-infected CD4+ T cells (Molt/LAV cells) were amplified and detected in infected individual cells using polymerase chain reaction and in rifu hybridization. In this in situ PCR, three parameters were considered to achieve effective amplification and retention of amplificants inside the cells by making high molecular weight PCR products intracellularly, forming agarose matrix against the cells, and maintaining the appropriate PCR temperature profile. Over the cycles of ampliHcationl tailed primers with complementary overhanging sequences at their 5' sides manufactured high molecular weight products by using short primary products as a repeating unit. Agarose matrix could prevent the diffusion of the amplificants from the cells. Use of Thermanox coverslip inside the PCR tube offered target cells a similar temperature profile to that of conventional PCR in solution.

간흡충 tropomyosin: PCR로 일부분 증폭된 cDNA의 cloning 및 염기서열 (Clonorchis sinensis tropomyosin: Cloning and sequence of partial cDNA amplified by PCR)

  • 홍성종
    • Parasites, Hosts and Diseases
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    • 제31권3호
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    • pp.285-292
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    • 1993
  • 간흡충 total RNk에는 많은 량의 185 rRNA가 함유되어 있었지만 285 rRNA는 그 양이 매우 적었다. 약 $8{\;}{\mu\textrm{g}}의{\;}poly{\;}(A)^{+}$ mRNAS부터 합성된 double-stranded CDNA는 대부분이 0.4-4.2 kb 크기이었으며 9.5 kb에 달하는 것도 있었다. 이미 보고되어 있는 tropomyosin의 amino산 서열을 기준하여 5개의 degenerated oligonucleotide (sense primer 2개와 antisense primer 3개)를 합성하였다. TotalcDNA를 template로 하고 sense primer와 antisense primer를 조합하여 실시한 PCR 산물 중에서 580 bp 크기의 특이 유전자가 나타났다. 만손주혈흡충의 tropomyosin CDNA를 탐색자로 써서 Southern hybridization했을 때 이 유전자만이 검출되어서. 이 유전자는 간횹충 tropomyosin (CSTM) CDNA의 일부분일 가능성이 높다고 생각되어 sequencing vector인 POEM-3Zf(-)에 cloning한 다음 염기서열을 결정하였다. nRf 증폭된 CSTM CDNA는 크기가 575 bp이었으며 191개의 predicted amino산 서열은 한 개의 open reading frame을 갖고 있었다 CSTM CDNA의 amino산 서열은 만손주혈흡충 tropomyosln과 86.3%. Trichosoonvk: colhnfornis tropomyosin과 51.1% 의 유사성을 갖고 있었다. 이 CSTM cDNA fragment는 앞으로 간흡충 cDNA library를 screening하여 완전한 CnM CDNA를 cloning하기에 좋은 probe로 쓰일 것으로 예상된다.

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