KR910004814A - 하플로타입으로서의 근 및 유전자 좌 대립 유전자의 검출을 위한 인트론 서열 분석 방법 - Google Patents

하플로타입으로서의 근 및 유전자 좌 대립 유전자의 검출을 위한 인트론 서열 분석 방법 Download PDF

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KR910004814A
KR910004814A KR1019900013222A KR900013222A KR910004814A KR 910004814 A KR910004814 A KR 910004814A KR 1019900013222 A KR1019900013222 A KR 1019900013222A KR 900013222 A KR900013222 A KR 900013222A KR 910004814 A KR910004814 A KR 910004814A
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제이. 시몬스 말콤
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Abstract

내용 없음.

Description

하플로타입으로서의 근 및 유전자 좌 대립 유전자의 검출을 위한 인트론 서열 분석 방법
본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음

Claims (34)

  1. 유전자 좌의 유전적 연쇄에 있으며 충분한 수의 인트론 서열 뉴클레오티드를 함유하고 있는 DNA 서열을 한정하는 인트론 스패닝 프라이머 쌍으로 게노믹 DNA를 확장시켜 대립 유전자의 특징적인 확장된 DNA 서열을 생성하는 것을 특징으로 하는, 유전자 좌의 적어도 하나의 대립 유전자의 검출방법.
  2. 제1항에 있어서, 확장된 DNA 서열이 인트론 서열에 해당하는 적어도 약 300 뉴클레오티드를 함유함을 특징으로 하는 방법.
  3. 제1항에 있어서, 인트론 서열이 대립 유전자를 코오딩하는 엑손에 인접함을 특징으로 하는 방법.
  4. 제1항에 있어서, 확장된 DNA 서열이 적어도 하나의 비인접 대립 유전자의 특징임을 특징으로 하는 방법.
  5. 제1항에 있어서, 확장된 DNA 서열이 적어도 하나의 인접 대립 유전자 및 적어도 하나의 비인접 대립 유전자의 특징임을 특징으로 하는 방법.
  6. 제5항에 있어서, 확장된 DNA 서열이 인트론 서열에 해당하는 적어도 약 1,000 뉴클레오티드를 함유함을 특징으로 하는 방법.
  7. a.대립 유전자와 유전적 연쇄에 있으며 충분한 수의 인트론 서열 뉴클레이티드를 함유하고 있는 DNA 서열을 한정하는 인트론 스패닝 프라이머 쌍으로 게노믹 DNA를 확장시켜 대립 유전자의 특징적인 확장된 DNA 서열을 생성시키고, b.확장된 DNA 서열을 분석하여 확장된 서열내의 유전적 변종의 존재를 검출함을 특징으로 하는, 적어도 하나의 대립 유전자 좌의 검출 방법.
  8. 제7항에 있어서, 확장된 DNA 서열내의 변종이 프라이머 한정 확장된 DNA 서열의 길이가 변종임을 특징으로 하는 방법.
  9. 제7항에 있어서, 확장된 DNA 서열내의 변종이 프라이머 한정 확장된 DNA 서열의 적어도 하나의 제한 부위가 변화됨을 특징으로하는 방법.
  10. 제7항에 있어서, 확장된 DNA 서열내의 변종이 프라이머 한정 확장된 DNA 서열의 적어도 하나의 제한 부위가 위치 변화됨을 특징으로하는 방법.
  11. 제7항에 있어서, 확장된 DNA 서열내의 변종이 프라이머 한정 확장된 DNA 서열의 적어도 하나의 뉴클레이티드가 치환됨을 특징으로하는 방법.
  12. 제7항에 있어서, 유전자 좌가 주요 조직적 합성 유전자 좌임을 특징으로 하는 방법.
  13. 제7항에 있어서, 대립 유전자가 일유전자성 질병과 관련됨을 특징으로 하는 방법.
  14. 제13항에 있어서, 일유전자성 질병이 낭포성 섬유증임을 특징으로 하는 방법.
  15. 제13항에 있어서, 적어도 약 70%의 프라이머 한정 확장된 DNA 서열이 인트론 서열에 해당함을 특징으로 하는 방법.
  16. 제7항에 있어서, 프라이머 한정 확장된 DNA 서열 길이가 300 내지 500 뉴클레오티드임을 특징으로 하는 방법.
  17. a.대상인의 게노믹 HLA DNA를, 서열을 생산하는데 적당한 조건하에서 HLA 유전자 좌에 특이적인 프라이머 쌍으로 확장시키고, b.확작된 DNA 서열을 절단하는 적어도 하나의 엔도뉴클레아제와 확장된 DNA 서열을 혼합하여 구별되는 단편 길이를 갖는 단편의 세트를 수득함으로써 절단된 혼합물을 생성하는 단계를 포함함을 특징으로하는, 대상인의 HLA 유전자 좌에 대한 RFLP 단편의 제조 방법.
  18. 제17항에 있어서, 절단된 혼합물로부터 RFLP 패턴을 생성하는 단계를 더 포함함을 특징으로 하는 방법.
  19. 제17항에 있어서, 프라이머가 HLA 유전자 좌와 관련된 대립 유전자 변종을 코오딩하는 모든 엑손을 포함하는 DNA 서열을 한정함을 특징으로 하는 방법.
  20. a.대상인의 게노믹 HLA DNA를, 확장된 DNA 서열을 생산하는데 적당한 조건하에서 HLA 유전자 좌와 관련된 대립 유전자 변종을 코오딩하는 모든 엑손을 포함하는 DNA 서열을 한정하는, HLA 유전자 좌에 특징적인 프라이머 쌍으로 확장시키고; b.확장된 DNA 서열을 절단하는 적어도 하나의 엔도뉴클레아제와 확장된 DNA 서열을 혼합하여 구별되는 단편 길이를 갖는 단편의 세트를 수득함으로써 절단된 혼합물을 생성하는 단계를 포함함을 특징으로 하는, 대상인의 HLA 유전자 좌에 대한 RELP 단편의 제조방법.
  21. a. 대상인의 HLA DNA를, 확장된 DNA 서열을 생산하는데 적당한 조건하에서 HLA 유전자 좌가 클래스 I 유전자 좌일 때 개재 서열 I 및 개재 서열 III에 위치하고, HLA 유전자 좌가 클래스 II 유전자 좌일때 개재 서열 I 및 개재 서열 Ⅱ에 위치하는 HLA 유전자 좌에 특이적인 프라이머 쌍으로 확장시키고; b. 확장된 DNA 서열을 절단하는 적어도 하나의 엔도뉴클레아제와 확장된 DNA 서열을 혼합하여 구별되는 단편 길이를 갖는 단편이 세트를 수득함으로써 절단된 혼합물을 생성하고; c. 절단된 혼합물로 부터 RELP 패턴을 생성하는 단계를 포함함을 특징으로 하는, 대상인의 HLA 유전자 좌에 대한 RELP 패턴의 생성 방법.
  22. 제22항에 있어서, 확장이 a.HLA 유전자 좌에 특이성 프라이머 쌍을, 프라이머 쌍의 각 프라이머가 그의 보체로 부터 분리될때, 다른 프라이머의 연장 생성물의 합성에 주형으로 작용할 수 있는 연장 생성물을 생성하기에 충분한 시간 동안 하이브리다아즈조건하에서 대상인의 HLA DNA와 혼합하여 혼합물을 제조하고; b.혼합물을 변성 조건하에서 처리하여 그의 연장 생성물로 부터 프라이머를 분리하고; c.혼합물을 프라이머 연장 생성물이 주형으로서 단계(b)에서 제조된 각 주형을 사용하여 합성되도록 HLA 유전자 좌 특이성 프라이머 쌍으로 처리하여 HLA DNA를 확장시키고; d. 단계(b) 및 (c)를 반복하여 확장된 DNA 서열을 생성하는 것으로 이루어짐을 특징으로 하는 방법.
  23. 제21항에 있어서, HLA 유전자 좌에 특이적인 제2프라이머 쌍을 또한 HLA DNA를 확장시키는데 사용함을 특징으로 하는 방법.
  24. 제21항에 있어서, RFLP 단편 패턴의 생성이 a.확장된 DNA 서열을 이 확장된 서열을 절단하는 적어도 하나의 엔도뉴클레아제와의 혼합하에서 구별되는 단편 길이를 갖는 단편의 세트를 수득하고; b.단편의 길이를 기초로 단편을 분리하여 분리된 단편을 제조하고; c.분리된 단편을 가시화하여 RFLP 단편 패턴을 생성하는 것으로 이루어짐을 특징으로 하는 방법.
  25. 제24항에 있어서, 단편을 겔 전기 영동을 사용하여 분리하고, 뉴클레오티드 특이성 염색을 사용하여 가시화함을 특징으로 하는 방법.
  26. a.대상인으로 부터의 DNA 및 시료로 부터의 DNA를, 대상인 및 시료로부터 확장된 DNA 서열을 생성하기에 적당한 조건하에서 HLA 클래스 I 유전자 좌의 경우에는 개개 서열 I 및 Ⅱ에 위치하고 클래스 III 유전자 좌의 경우에는 개재 서열 I 및 Ⅱ에 위치하는, HLA 유전자 좌에 특이적인 프라이머 쌍으로 확장시키고; b.대상인으로 부터의 확장된 DNA 서열 및 시료로 부터의 확장된 시료 DNA를, 확장된 DNA 서열을 확장된 DNA의 분해에 충분한 시간 동안 HLA 유전자 좌의 대립 유전자들을 구별하기에 충분히 다른 길이의 다수의 절단된 서열로 절단하는 적어도 하나의 엔도뉴클레아제와 혼합하여 절단된 혼합물을 생성하고; c.대상인 및 시료로 부터의 절단된 혼합물에 의해 생성된 제한 단편 길이 다형 패턴을 비교하는 단계를 포함함을 특징으로 하는, 시료 중의 DNA가 특정 대상인의 것인지의 여부를 결정하는 방법.
  27. a.대상인으로부터의 DNA, 어린이로 부터의 DNA 및 어린이의 어머니로부터의 DNA를, 확장된 DNA 서열을 생성하는데 적당한 조건하에서 HLA 클래스 I 유전자 좌의 경우 개재 서열 I 및 Ⅱ에 위치하고, 클래스 Ⅱ 유전자 좌의 경우 개재 서열 I 및 Ⅱ에 위치하는, HLA 유전자 좌에 특이적인 프라이머의 쌍으로 확장시키고; b. 대상인으로부터의 확장된 DNA 서열 및 어린이로부터의 확장된 DNA 서열, 확장된 DNA 서열을 HLA 유전자 좌의 대립 유전자들을 구별하기에 충분히 다른 길이의 다수의 절단된 서열도 절단하는 적어도 하나의 엔도뉴클레아제와 혼합하여 절단된 혼합물을 생성하고; c.대상인, 어린이의 어머니 및 어린이로 부터의 절단된 혼합물에 의해 생성된 제한 단편 길이 다형 패턴을 비교하는 단계를 포함함을 특징으로 하는, 대상인이 어린이의 아버지인지의 여부를 결정하는 방법.
  28. 클래스 I 유전자 좌 특이성 프라이머, 클래스 I A 유전자 좌 특이성 프라이머, 클래스 IC 유전자 좌 특이성 프라이머로 구성된 군으로부터 선택된 HLA 유전자 좌 특이성 프라이머.
  29. 제28항에 있어서, 프라이머가 CATGTGGCCATCTTGAGAATGGA;
    의 군에서 선택된 적어도 15연속 뉴틀레오티드에 해당하는 서열을 가짐을 특징으로 하는 HLA 유전자 좌 특이성 프라이머.
  30. HLA 클래스 I 유전자 좌 특이성 프라이머 쌍.
  31. HLA 클래스 Ⅱ 유전자 좌 특이성 인트론 스패닝 프라이머 쌍.
  32. HLA 유전자 좌 특이성 프라이머 쌓에 의해 한정된 DNA 서열.
  33. 적당한 용기 내에 HLA 클래스 I 유전자 좌 특이성 프라이머 쌍 및 HLA 클래스 Ⅱ 유전자 좌 특이성, 인트로 스페닝 프라이머 쌍의 군에서 선택된 적어도 하나의 HLA 유전자 좌 특이성 프라이머 쌍을 포함함을 특징으로 하는 키트.
  34. 제33항에 있어서, HLA 유전자 좌 특이성 프라이머 쌍에 의해 한정된 DNA 서열을, HLA 유전자 좌의 대립 유전자들을 구별하기에 충분히 다른 길이의 절단된 서열도 절단하는 적어도 하나의 엔도 뉴클레아제를 더 포함함을 특징으로 하는 키트.
    ※ 참고사항 : 최초출원 내용에 의하여 공개하는 것임.
KR1019900013222A 1989-08-25 1990-08-24 하플로타입으로서의 근 및 유전자 좌 대립 유전자의 검출을 위한 인트론 서열 분석 방법 KR910004814A (ko)

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