GI I TRÌNH T GENẢ Ự
C¸c ph¬ng ph¸p gi¶i tr×nh tù gen
Cã 2 ph¬ng ph¸p chÝnh:
–Ph¬ng ph¸p ho¸ häc Maxam – Gilbert: ®©y lµ ph
¬ng ph¸p ®îc sö dông ®Çu tiªn ®Ó x¸c ®Þnh
tr×nh tù gen nhng hiÖn nay kh«ng cßn ®ù¬c sö
dông phæ biÕn n÷a.
–Ph¬ng ph¸p Enzyme Sanger
1. Ph¬ng ph¸p ho¸ häc Maxam Gilbert
Walter Gilbert
Nguyªn lý
Sö dông c¸c chÊt ho¸ häc c¾t ®øt sîi DNA t¹i 1 lo¹i
nucleotide ®Æc hiÖu
Ph©n tö DNA kÐp ®îc ®¸nh dÊu P32 t¹i ®Çu 5’
T¸ch rêi 2 sîi ®¬n b»ng nhiÖt
Xö lý ho¸ häc sao cho chØ cã 1 lo¹i nucleotide bÞ
ph¸ huû -> hµng lo¹t ®o¹n DNA cã kÝch thíc kh¸c
nhau ®îc t¹o thµnh (sö dông c¸c chÊt ho¸ häc kh¸c
nhau ph¸ huû 4 lo¹i nucleotide b»ng 4 ph¶n øng riªng
biÖt)
TiÕn hµnh ®iÖn di c¸c ®o¹n DNA thu ®ù¬c trªn gel
polyacrylamide, chØ quan s¸t ®ù¬c ®o¹n cã g¾n P32.
§o¹n DNA ng¾n nhÊt cã ®¸nh dÊu phãng x¹ sÏ ch¹y
nhanh nhÊt díi t¸c dông cña ®iÖn trêng.
C¨n cø vµo vÞ trÝ c¸c v¹ch quan s¸t ®îc trªn gel mµ
x¸c ®Þnh tr×nh tù c¸c nucleotide.
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