Biomedical Engineering Reference
In-Depth Information
Cla
I
-
Bsp
D
I
6881
Swa
I
6782
Acl
I
6772
Dsa
I
6616
BsmF
I
6528
Bpm
I
6904
Bgl
II
6934
Bsu
36
I
6507
Bgl
I
6431
Fsp
I
6425
Pvu
I
6405
Ear
I
6390
Bpm
I
6492
Msl
I
6996
Xmn
I
357
Polylinker
cloning sites
II
6229-6287
Bsa
H
I
6000
I
-
Kas
I
6000
Bsr
G
I
1020
Bsa
B
I
1148
Sna
B
I
1267
Bsp
H
I
1298
Sfo
I
-
Nar
Bsm
B
I
5970
Ava
II
5914
Drd
I
5758
Dra
III
5715
Bso
B
I
-
Ava
I
5824
Plus
ORI
Minus
Ban
II
5642
Nae
I
-
Ngo
M
IV
5612
Ban
II
5642
Bsr
F
I
5612
Bsm
I
1745
Bsm
F
I
5089
Bse
R
I
2007
Alw
N
I
2186
Msc
I
5079
Cla
I
-
Bsp
D
I
2526
Xmn
I
2645
Dsa
I
2762
Msl
I
2795
Acl
I
4633
I
Pac
I
4131
Eco
47
III
3038
Bsa
B
I
3973
Ear
I
4073
1
(Met) Thr Met Ile Thr Asn Ser Ser Ser Val Pro Gly Asp Pro Leu Glu Ser Thr Cys Arg His Ala Ser Leu Ala
→
LacZ´
6230
6250
6270
6290
Ecl
136
II
Sac
I
Xma
I
Sma
I
Xba
I
Pst
I
Hin
d
III
atgaccatgatta
cgaattC
GAGCTC
GGTACC
CGG
GGATCC
TCTAGA
GTCGAC
CTGCAG
GCATGC
AAGCTTGGcact
Eco
R
I
Kpn
I
Acc
65
I
Bam
H
I
Sal
I
Hin
c
II
Acc
I
Sph
I
Ava
I
/
Bso
B
I
Fig. 4.17
Restriction map of cloning vector M13mp18. Phages M13mp18 and M13mp19 are 7249 bases in length and differ
only in the orientation of the 54-base polylinker that they carry. The map shows the restriction sites of enzymes that cut the
molecule once or twice. The unique sites are shown in bold type.
making detection of recombinants easy. However,
the
lac
region only contains unique sites for
Ava
II,
Bgl
I and
Pvu
I and three sites for
Pvu
II, and there are
no sites anywhere on the complete genome for the
commonly used enzymes such as
Eco
RI or
Hin
dIII.
To remedy this defect, Gronenborn and Messing
(1978) used
in vitro
mutagenesis to change a single
base pair, thereby creating a unique
Eco
RI site within
the
lac
fragment. This variant was designated M13
mp2. This phage derivative was further modified to
generate derivatives with polylinkers upstream of
the lac
-fragment (Fig. 4.17). These derivatives
(mp7-mp11, mp18, mp19) are the exact M13
counterparts of the pUC plasmids shown in Fig. 4.9.
α
