KR890013184A - 핵산을 증폭시키는 방법 - Google Patents

핵산을 증폭시키는 방법 Download PDF

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KR890013184A
KR890013184A KR1019890002142A KR890002142A KR890013184A KR 890013184 A KR890013184 A KR 890013184A KR 1019890002142 A KR1019890002142 A KR 1019890002142A KR 890002142 A KR890002142 A KR 890002142A KR 890013184 A KR890013184 A KR 890013184A
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primer
dna
nucleic acid
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다베이 체릴
티이.말렉 로렌스
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로버트 티이.가아빈
칸진 코포레이션
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    • C—CHEMISTRY; METALLURGY
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    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
    • C12Q1/701—Specific hybridization probes
    • C12Q1/702—Specific hybridization probes for retroviruses
    • C12Q1/703—Viruses associated with AIDS
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6844—Nucleic acid amplification reactions
    • C12Q1/6865—Promoter-based amplification, e.g. nucleic acid sequence amplification [NASBA], self-sustained sequence replication [3SR] or transcription-based amplification system [TAS]
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C12Q1/689—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for bacteria

Abstract

내용 없음.

Description

핵산을 증폭시키는 방법
본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음
제1도는 핵산 증폭 방법을 일반적으로 설명한 것이다,
제2도는 증폭 방법을 검사하는데 사용하는 합성 올리고 뉴클레오티드 DNA서열을 나타낸다 : 제2도 A, gag test서열 ; 제2도 B, gag2 test서열,
제3도는 다른 프라이머 농도를 사용한 증폭 반응을 PAGE분석한 자동방사선 사진이다.

Claims (42)

  1. 제1프라이머에 대한 제1주형인 한 가닥 RNA, 제2프라이머에 대한 제2주형인 한 가닥 DNA 및 많은 복제수의 제1주형을 합성하기 위한 제3주형인 이중 가닥 DNA를 합성하는 것으로 되고, 여기에서, 제1또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되는 것이 특징인 특이한 핵산 서열을 증폭시키는 방법.
  2. 제1항에 있어서, 제1프라이머가 선택적으로 제1주형에 혼성화하고 제1프라이머에 공유 결합된 DNA의 합성을 촉진하며 제1주형에 상보적이고, 프로모터에 대한 안티센스 서열 및 RNA폴리머라제의 전사 개시 부위에 대한 안티센스 서열을 갖는 제2프라이머가 제2주형에 선택적으로 혼성화함으로서 제2프라이머에 공유 결합된 DNA의 합성을 가능하게 하고 제2주형에 상보적인 것이 특징인 방법.
  3. (a) 제1주형의 RNA서열에 대해 충분히 상보적인 서열을 갖는 DNA의 제1프라이머를 제1주형에 혼성화하고, (b) 제1프라이머에 공유 결합되고 제1주형의 RNA서열에 상보적이며, 제1프라이머와 함께 제2주형을 구성하는 제1DNA서열을 합성하고, (c) 제2프라이머를 혼성화하도록 제2주형으로 부터 제1주형을 분리하고, (d) 제2주형의 DNA서열에 대해 충분히 상보적인 서열을 가지며 프로모터의 안티센스 서열 및 RNA폴리머라제의 전사 개시부위에 대한 안티센스 서열을 갖는 DNA의 제2프라이머를 제2주형에 혼성화하 고, (e) 제2프라이머에 공유 결합되고 제2주형의 DNA서열에 상보적인 제2DNA서열과, 제2주형에 공유 결합되고 제2프라이머의 DNA서열에 상보적인 제3DNA서열을 합성하여, 상기 제2 및 제3DNA서열, 제2프라이머 및 제2주형으로 구성되는 제3주형을 얻고, (f) 제3주형으로부터 다수의 복제된 제1주형의 RNA 서열을 합성하는 것으로 되고, 여기에서, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대한 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되어 있는 것이 특징인 특이한 핵산 서열을 증폭시키는 방법.
  4. 제3항에 있어서, 제2프라이머가 그의 3'말단에 제2주형의 DNA서열에 대해 충분히 상보적인 서열을 가지며, 그의 5'말단에 프로모터의 안티센스 DNA서열 및 RNA폴리머라제에 대한 전사 개시 부위의 안티센스 서열을 갖는 것이 특징인 방법.
  5. 제4항에 있어서, 제2주형에 공유 결합된 제3DNA서열이 제2프라이머의 5'말단에서 DNA서열에 대해 상보적인 것이 특징인 방법.
  6. 제3항에 있어서, 제1DNA서열이 RNA의존성 DNA폴리머라제에 의해 합성되는 것이 특징인 방법.
  7. 제3항에 있어서, 제1DNA주형이 RNA-DNA혼성체에 대해 특이성을 갖는 리보뉴클레아제에 의해 제2주형으로 부터 분리되는 것이 특징인 방법.
  8. 제7항에 있어서, 리보뉴클레아제가 E.coli 리보뉴클레아제 H인 방법.
  9. 7항에 있어서, 리보뉴클레아제가 소의 흉선 리보뉴클레아제 H인 방법.
  10. 제3항에 있어서, 제2 및 제3DNA서열이 DNA의존성 DNA폴리머라제에 의해 합성되는 것이 특징인 방법.
  11. 제3항에 있어서, 제1주형이 DNA의존성 RNA폴리머라제에 의해 제3주형으로 부터 합성되는 것이 특징인 방법.
  12. 제3항에 있어서, 제1프라이머가 프로모터의 안티센스 서열 및 전사 개시 부위의 안티센스 서열을 갖는 것이 특징인 방법.
  13. 제3항에 있어서, 제1프라이머가 지지 수단에 혼성화하는 서열을 갖는 것이 특징인 방법.
  14. 제3항에 있어서, 제2프라이머가 그의 3'말단에서 부터 5'말단 사이에 충분히 상보적인 서열, 프로모터의 안티센스 서열 및 전사 개시 부위의 안티센스 서열을 갖는 것이 특징인 방법.
  15. 제14항에 있어서, 전사 개시 부위의 안티센스 서열 및 안티센스 프로머터 서열이 박테리오파지 RNA폴리머라제와 결합하는 것이 특징인 방법.
  16. 제14항에 있어서, 안티센스 프로모터 서열 및 전사 개시 부위의 안티센스 서열이 T7 RNA폴리머라제와 결합하는 것이 특징인 방법.
  17. 제14항에 있어서, 안티센스 프로모터 서열 및 전사 개시 부위의 안티센스 서열이 AATTCTAATACGACTCACTATAGGGAG인 것이 특징인 방법.
  18. 제11항에 있어서, DNA의 의존성 RNA폴리머라제가 T7 RNA폴리머라제인 것이 특징인 방법.
  19. 제11항에 있어서, DNA의존성 RNA폴리머라제가 박테리오파지 RNA폴리머라제인 것이 특징인 방법.
  20. 제11항에 있어서, DNA의존성 RNA폴리머라제가 파지 T3폴리머라제인 것이 특징인 방법.
  21. 제11항에 있어서, DNA의존성 RNA폴리머라제가 파지 ø11폴리머라제인 것이 특징인 방법.
  22. 제11항에 있어서, DNA의존성 RNA폴리머라제가 살모넬라 파지 sp6폴리머라제인 것이 특징인 방법.
  23. 제11항에 있어서, DNA의존성 RNA폴리머라제가 슈도모나스 파지 gh-1폴리머라제인 것이 특징인 방법.
  24. 제6항에 있어서, RNA의존성 DNA폴리머라제가 레트로바이러스의 폴리머라제인 것이 특징인 방법.
  25. 제24항에 있어서, 레트로바이러스의 폴리머라제가 조류의 근원세포 분해 바이러스의 폴리머라제인 것이 특징인 방법.
  26. 제24항에 있어서, 레트로바이러스의 폴리머라제가 말로니 쥐의 백혈구 바이러스의 폴리머라제인 것이 특징인 방법.
  27. 제6항, 제10항 또는 제11항에 있어서, 폴리머라제가 엑소뉴클레아제 또는 엔도뉴클레아제 활성이 결핍된 것이 특징인 방법.
  28. 제10항에 있어서, DNA의존성 DNA폴리머라제가 조류의 근원세포 분해 바이러스의 폴리머라제인 것이 특징인 방법.
  29. RNA의 제1주형에 대해 충분히 상보적인 서열을 가지는 DNA의 제1프라이머, 제2주형, 프로모터의 안티센스 서열 및 RNA폴리머라제에 의해 기질로서 인식되는 전사 개시 부위의 안티센스 서열에 충분히 상보적인 서열을 갖는 DNA의 제2프라이머, 리보뉴클레아제 H, RNA-의존성 DNA폴리머라제, DNA의존성 DNA폴리머라제, RNA폴리머라제, 리보뉴클레오시드 트리포스페이트 및 데옥시리보뉴클레오시드 트리포스페이트를 시료와 결합시키는 것으로 되고, 제1프라이머 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되는 것이 특징인 특이한 핵산 서열의 증폭방법.
  30. RNA의 제1주형에 대해 충분히 상보적인 서열을 갖는 제1프라이머, DNA의 제2주형, 프로모터의 안티센스 서열 및 RNA폴리머라제에 의해 기질로서 인식되는 전사 개시 부위의 안티센스 서열에 충분히 상보적인 서열을 갖는 제2프라이머, 조류의 근원세포 분해 바이러스의 폴리머라제, 대장균 리보뉴클레아제 H, 박테리오파지 T7 DNA폴리머라제, 리보뉴클레오시드 트리포스페이트 및 데옥시리보뉴클레오시드 트리포스페이트를 시료와 결합시키는 것으로 되고, 제1프라이머 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되는 것이 특징인 핵산 을 증폭시키는 방법.
  31. 제1항 또는 제3항에 있어서, 추가로 핵산 서열을 함유한 것으로 예견되는 제1시료의 증폭량과 핵산 서열이 존재하지 않는 제2시료의 증폭량을 비교함으로서 특이한 핵산 서열의 존재를 탐지하는 것을 포함하는 방법.
  32. 제29항 또는 제30항에 있어서, 추가로 핵산 서열을 함유한 것으로 예견되는 제1시료의 증폭량과 핵산 서열이 존재하지 않은 제2서열의 증폭량을 비교함으로서 특이한 핵산 서열의 존재를 탐지하는 것을 포함하는 방법.
  33. 제1항 또는 제3항에 있어서, 추가로 특이한 핵산의 존재를 특이한 핵산 서열 또는 특이한 핵산 서열의 산물과 혼성화하는 프로브를 탐지하는 것을 포함하는 방법.
  34. 제1항 또는 제3항에 있어서, 추가로 제한 효소 및 전기영동 분리법을 사용하여 특이한 핵산 서열의 존재를 탐지하는 것을 포함하는 방법.
  35. 제31항 또는 제32항에 있어서, 추가로 제한 효소 및 전기영동 분리법을 사용하여 특이한 핵산 서열의존재를 탐지하는 것을 포함하는 방법.
  36. 제31항 또는 제32항에 있어서, 추가로 특이한 핵산 서열을 특이한 핵산 서열 또는 특이한 핵산 서열의산물 혼성화시킨 프로브로 탐지하는 것을 포함하는 방법.
  37. 제1항 기재의 방법에 의해 증폭된 특이한 핵산 서열.
  38. 제3항 기재의 방법에 의해 증폭된 특이한 핵산 서열.
  39. 제29항 기재의 방법에 의해 증폭된 특이한 핵산 서열.
  40. 제30항 기재의 방법에 의해 증폭된 특이한 핵산 서열.
  41. (a) 제1주형의 RNA서열에 충분히 상보적인 DNA서열을 갖는 제1프라이머, (b) 제2주형의 DNA서열에 상보적인 DNA서열을 갖는 제2프라이머, (c) 리보뉴클레아제 H, (d) RNA-의존성 DNA폴리머라제, (e) DNA-의존성 RNA폴리머라제, (f) DNA-의존성 DNA폴리머라제, (g) 리보뉴클레오시드 트리포 스페이트 및 (h) 데옥시리보뉴클레오시드 트리포스페이트로 구성되고, 상기 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되는 것이 특징인 특이한 핵산 서열을 증폭시키기 위한 키트(kit).
  42. (a) 제1주형의 RNA서열에 충분히 상보적인 DNA서열을 갖는 제1프라이머, (b) 제2주형의 DNA서열에 충분히 상보적인 DNA서열을 갖는 제2프라이머, (c) 박테리오파지 T7 ANA폴리머라제, (d) 대장균 리보뉴클레아제 H, (e) 조류의 근원세포 분해 바이러스의 폴리머라제, (f) 리보뉴클레오시드 트리포스페이트 및 (g) 데옥시리보뉴클레오시드 트리포스페이트로 구성되고, 상기 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분히 상보적이고, 제1 또는 제2프라이머의 서열이 특이한 핵산 서열의 서열에 대해 충분한 상동성을 가지며, 제1프라이머의 3'말단이 상보성 가닥위의 제2프라이머의 3'말단을 향해 배향되는 것이 특징인 특이한 핵산 서열을 증폭시키기 위한 키트(kit).
    ※ 참고사항 : 최초출원 내용에 의하여 공개하는 것임.
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DE3850093T2 (de) 1994-11-03
ES2053648T3 (es) 1994-08-01
KR920702866A (ko) 1992-10-28
US5409818A (en) 1995-04-25
WO1991002814A1 (en) 1991-03-07
ATE106948T1 (de) 1994-06-15
EP0329822A2 (en) 1989-08-30
EP0329822B1 (en) 1994-06-08
EP0329822A3 (en) 1990-08-01
DE3850093D1 (de) 1994-07-14
KR960015744B1 (ko) 1996-11-20
CA1340807C (en) 1999-11-02

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