??? ?????PCR-RFLP??

1
??? ?????PCR-RFLP??
???????
2
?????? 1?????DNA??? 2??????PCR?? 3??????PCR?? 4??
??????(?????)
3
?DNA??? 1??????????2EDTA?? 2?DNA??
?????(??????) ?-??????? ??RNA ??DNA
4
PCR?? 1?????(Genbank) 2???????(polymerase
chain reaction,PCR)
5
PCR???? DNA?? dNTP Taq DNA??? ?? Mg2
???
6
PCR???? ??? 94? 35min 35?? ??
94 ? 30s1min ?? 5065 ?
3060s ?? 72 ? 12min
??? 72 ? 510min
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PCR????????
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?????PCR??
PCR???????? 5-TCATGA-3
3-AGTACT-5
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??????(1)
A
B
????
AA
BB AB
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??????(2)
11
?????? ????????????
12
??????

• ????DNA??????????pH???????,??????????
• ???????????????,??DNA???DNA????????,?????????????
  
• ???????????

13
??????

• ???????,????????????DNA??????????????

14
????????????

• ??DNA??
• ????????????????????????Tip?
• ??
• ?????(1TAE)0.04mol/L????????-??(tris-??),0.01mol
  /L????????(EDTA)
• ????(EB10mg/ml)100ml????????1g EB
• 1???100ml 1TAE??1g???
• ?????(???-????)???1????????????????

15
??????
16
(?)??

• 1.?1.0g?????100 ml 1TAE????,??1?????????????????
  ??????????60?C???
• 2.??5ul????(???0.5ug/ ml )???????????,????????
• 3.?????????
• 4.???????????????,?????
• 5.??????????,???????????,?????????????,?????????
• 6.????,???????????(?20??),?????????2L
  1TAE???,???????????????????

17
(?)??

• 1.????????????????
• 2.??????????,???????????1cm?
• 3.?5ul?????????,??????????????
• 4.????,??4-5V/cm,DNA??????????30-40???
• 5.????,?????

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??????
1. ???????????????????????? 2.?????,???????? 3.??
?????,??????????? 4.????????????????
19
????

• 1.??????
• 2.????
• 3.????????MspI???????6???(??),???6????,????????3
  ?????A?B?C??,???16????