??? DNA????????

1
??? DNA????????
2
?????????? ??? ?????????? ??? ???? ???
?????
??? ???? ??? ????(????) ??? ????????
3
??????? ? ?????(??????????????????
?????? ????????DNA???) ? ?????????(????????)
? ?????????(?????????) ???????
???????(???)???????(????
?)??????????????????(???? ?)????????
4
??????? ? DNA????????(?????) ?
???-N-??????? (???U????C?????U) ?
?????? ? ??????(?????????SOS???)
? ??????-----????DNA???
5
??? ???????????
6
1??????????????????? ?????????102103?
??????(MRS Mismatch Repair System)
DNApol (? 10-8) ??????? 10-11
7
2???????(Mismatch repair system)
(1)??
DNA polymerase Helicase SSB ????? (???)
???
MCE (mismatch correct enzyme) 3
subunits mutH, L, S
8
? DNA???????????
DNA??GATC(palindromic seq.) ?m6A???????
???2kb????GATC seq.
?????????????
9
(2) ????
a?MutH/MutS ?????? ??????GATC??
??--???GATC? G?5?
????????
DNA helicase II, SSB,
exonuclease I????? ??????
DNA polymerase III ? DNA ligase ????
?????????DNA????
10
???????????5?(????????5?) ----------------?----
------------3?(--------------------------3?)
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3? ???-N-????? ( ung system )
????????dUTP???
??????????
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---TAGC--- ---A CG---
G
C
U
A
U
---TAGC--- ---ATCG---
---TAGC--- ---A CG---
---TAGC--- ---ATCG---
13
??? DNA?????
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??????????
???????
???????
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(No Transcript)
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??????????
1????(photo reactivation )

• ?????????

• 400 nm ???PR ?
• (photo-reactivation enzyme)
• ?????(photolyase)

----TT---- ----AA----
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2. ???? ExisionRepair
??????

• ?????
• ????

???????

• UvrA, B, C gene
• ?????
• (Endonucleases)
• ?????
• (Exonuclease)
• DNA pol
• Ligase

18
?
?
???
?
Pol? ???
???
?
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3. ????(RecombinativeRepair)
??????E.coli?????
??????????
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? Rec-A. gene ???????DNA????
? Rec??????????????
???? ? Uvr????????? ?
Rec??????????????? ???????
21
? ???????
22
??????
? ?Rec-A???????(strand transfer)??
? TT dimer????,?????????TT dimer
?????
? ???????,?????????
23
???? (?????)

• ?????
• ????

??????

• RecA, DNApolymerae
• ligase

RecA
??????????? ??????????
???????
24
4. ????(SOS?? SOS repair)
(1) ????
? 80 10
100 mut. 100
50 10
UV ??? W??(Jean Weigh)

• ?????? DNA ?E.coli A???
• E. coli ?SOS????U.V.?? (A B)
• SOS ?????????????(???)

25
(2) SOS ????
SOS ??--??????DNA??
?????????,????????
SOS????,???????????????
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SOS ????SOS??????
RecA?SOS?? ????????
RecA?LexA?????
RecA?LexA??????
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RecA-P ????
28
?DNA????/ DNA damaged
29

• ???????
• 1???????????
• ??????
• ???-N-???????
• 2?DNA??????
• ?????
• ?????
• ????
• ????
• ????(SOS??)

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?DNA???????
31
??? ?????
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1? ??-????(restriction and modificaion)
50??? Luria(T?????) Bertani(?)
Weigle(P2???)
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E.coli K??? B????????????? E.coli C????
??
? ????????????????????DNA?? ?
?????DNA????????? ??????-????
???????-?????????DNA???DNA
34
??DNA ???????DNA
????????DNA????????????? ??DNA(resident
DNA)????????????DNA,
????DNA,??DNA,???DNA?
35
2???-????
??????????(???)
36
??? ????
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??(mutation)??????????DNA??
????????
????
? ?????(Chromosome mutation )
chromosome number chromosome structure
? ?????( dNt point mutation )
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???(mutant)???????????????
????(mutantgene) ?????????
?????(wild type gene)
???? ??? Arg Arg- ???
arg arg- ??? ???
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1??????????? ????--??????????????
?? --???????
? ???????DNA??????? -----????
?????
40
???????DNA?????
41
???????DNA?????
G(k)
C(i)
C(a)
C(i)
C(a, anti)
G(k, syn)
G(k, syn)
G(k, anti)
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2??????????? ????(point mutation) (???)
????(multiple mutation)
???---??????????????
? ????( conversion )
43
3????????????
????(frameshift mutation)??????????

???,?????????
????
? ???????? (dNt deletion or insertion )
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Examples of deletion mutations
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4?????????????(???) ???????????????????
??????????????????????
(?????????) ??????????????????????
???????????
? ????????????
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??????
UAA (Oc) ???(Ocher)
UAG(Am) ???(amber)
UGA(Opal) ???(Opal)
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5????????? ???
---??????
---?????
????? ????? ????
(?,?????)
6????????? ???? ?????????????????????????
?? ??????????? ????????????----????
48
7??????????????????
??????? ???????
???????????????? ???????
??????????? ????? ?????????????????
??????
???????????????? ??????????????????
?????????????????
49
??? ???? (????)
50
wild type
mutant (???)
1??????????
?????????????????????? ????????????????????????
???
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2????????????
?????? ??????????????????
????????????
?????? ?????????????????,?
??????????????????
????????????,???????
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3??????????
a) AGC (Ser) ? ACC (Thr) ? AGC (Ser)
b) AGC (Ser) ? AGG (Arg) ? AGT (Ser)
c) AGC (Ser) ? AGG (Arg) ? GGG (Gly)
if
Ser Gly
53
4??????????
(1)??????? (Intragenic suppression)
???? ????
???????????? ?????????????
??????????????--- ?????????????????
(?????????)
54
a?????
55
(No Transcript)
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----UGG 174-----------GGA210----
C
(C) (G)
----UGG174----------GGA210----
A
(K) (E)
????
----UGC174-----------GAA210----
57

• ???????
• 1????????
• 2??????
• ??????? ??????? ???????
• ?????????????????????
• ?????????????????
• 3? ????
• ???????????
• ???????????????????
• ??????????????

58
(No Transcript)
59
(2)????????
????---?????? ????---?????? ????---??????
??? tRNA????tRNA????????
60
a???????????( Nonsense suppressor )
???
UAG?UGA?UAA---?????? 50
???????tRNA???????,???????
61
b???????????( Missense suppressor )
62
c??????????
????????????(??)????????
??????(??)???????
63
??? ????????
64
1??????(Base analog)
2-???? 2-Amino purine
5-???? 5-Bromine Uracil
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G
A
AT?GC ??
5-BrU
?????? GC?AT ??
66
??????????????
67
2? ???????????
??????? ???(nitrous acid HNO2)
??(hydroxylamine HA) ?????(ethyl
mathanesulfonate EMS) N-??-N-??-N-????
(N-mathyl-N-nitro-N-nitrosoguanidion NNG)
??---????
68
NH2OH (Hydroxylamine HA ??)
69
3??????????
??? (Acridine Orange AO)
??????
???? (Ethidium Bromide EB )
????
-ATTTTTCG - -TAAAAAGC-
-AT EB TTTTCG- -TA
AAAAGC-
X
????---????
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4? ??????????
5? ????(mutator gene )
??????????????????????? ????,??????????????
????
71
??????
72
6??????????
73
??????????????????
74
7? ????(Mutation Hotpoint)
?????????????
DNA?????????????????,???????----
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a) ???? (repetitive seq. )/ palindromic seq.
b) m5C
???????????????, ???????????? ????????50
c) ???????????,?????????
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DNA ????????
Mutagen (???)
???????
DNA ??
?????
????
??
????
77

• ????
• ???? ???? ???? ???? ?????
• ??DNA ????? ????
• ???
• 1?????????????????????????
• 2?m5C??????????
• 3??????????????????
• 4?????????????????
• 5????-N-?????????????????
• 6????????????????????????
• 7???RecA??,??SOS????????
• 8?????????????????

78
????!!
79

• 2003-2004???? 6??