Restriction Enzyme Sensitivity to CpG Methylation
Methylation
Restriction Enzyme Properties
Many prokaryotes and eukaryotes contain enzymes known as DNA methyltransferases, which modify DNA by methylating cytosine in the CpG sequence. While these
enzymes are know to play a role in protecting host DNA from cleavage by corresponding restriction endonucleases in prokaryotes, their role in eukaryotes is not clear.
Enzyme These Takara enzymes
Site are listed
However, several restriction enzymes will be unable to cleave eukaryotic DNA at sites that have been methylated by CpG methylation.
PshA I
GACN GTC2
below. Methylation may occur at CpG sequences either within the recognition site (indicated by boldface color type), or overlapping it (indicated by boldface4 type).
Enzyme
Aat II
Acc I
Acc II
Afa I
Aor13H I
Aor51H I
Apa I
ApaL I
Ava II
Avi II
Bgl I
Bmg T1201
BspT104 I
BssH II
Bst1107 I
Cfr10 I
Site
GACGTC
GTMKAC
CGCG
GTAC
TCCGGA
AGCGCT
GGGCCC
GTGCAC
GGWCC
TGCGCA
GCCN5GGC
CCWGG
TTCGAA
GCGCGC
GTATACG
RCCGGY
Enzyme
Cfr13 I
Cla I
Cpo I
Eae I
Eco52 I
EcoR I
Fse I
Hae II
Hap II
Hha I
Hin1 I
Mlu I
Nae I
Nhe I
Not I
Nru I
PmaC I
Site
GGNCC
ATCGAT
CGGWCCG
YGGCCR1
CCGGCCG
GAATTCG
GGCCGGCC
RGCGCY
CCGG
GCGC
GRCGYC
ACGCGT
GCCGGC
GCTAGC
GCGGCCGC
TCGCGA
CACGTG
Psp1406 I
Pvu I
Sac
II
Enzyme
Sal
I I
PshA
Sau3A I
Psp1406 I
Sfi I
Pvu I
Sma
Sac III
SnaB
Sal I I
VpaK11B
Sau3A I I
Xho
Sfi II
AACGTT
CGATCG
CCGCGG
Site
GTCGAC
GACN GTC2
GATC 4
AACGTT
GGCCN5GGCC3
CGATCG
CCCGGG
CCGCGG
TACGTA
GTCGAC
GGWCCG
GATC
CTCGAG
GGCCN5GGCC3
Smawhen
I
CCCGGG
only
R=G
methylation
at multiple sites
required to
SnaB I
TACGTA
block
cleavage
VpaK11B
I
GGWCCG
3
internal
when the adjaXho I site methylated only
CTCGAG
cent N = G
1
only when R = G
2
methylation at multiple sites required to
block cleavage
3
internal site methylated only when the adjacent N = G
1
2
dcm Methylation
Restriction Enzyme Sensitivity to dam anddam
and dcm Methylation
DNA prepared from most common strains of E. coli may prove resistant to cleavage by certain restriction enzymes. This can often be caused by site-specific methylation. Two
types of methylases are commonly encountered: the dam methylase, which methylates the N6 position of the adenine residues in the sequence GATC, and the dcm methylase, which methylates the internal cytosine residues in the sequences CCWGG at the C5 position. The sensitivity of Takara’s enzymes to these types of methylation is listed
below.
DNA prepared from most common strains of E. coli may prove resistant to cleavage by certain restriction enzymes. This can often be caused by site-specific methylation. Two
types of methylases are commonly encountered: the dam methylase, which methylates the N6 position of the adenine residues in the sequence GATC, and the dcm methysensitivity
of Takara’s
enzymesdcm
to Methylation
these types of methylation is listed
lase, which methylates
the Blocked
internal by
cytosine
residuesdam
in the
sequences CCWGG at the C5 position. The
Enzymes
Overlapping
Methlyation
Enzymes
Blocked
by Overlapping
below.
Enzyme
Site
Enzyme
Site
Restriction Enzyme Sensitivity to
Acc III
Cla I
Fba I
Mbo I
Mbo II
Mfl I
Nru I
TthHB8 I
Xba I
TCCGGATC
ATCGAT(C)
TGATCA
GATC
GAAGA(TC)
RGATCY
TCGCGA(TC)
TCGA(TC)
TCTAGA(TC)
dam and dcm Methylation
Apa I
Ava II
Bal I
BmgT 1201
Eae I
EcoO109 I
Sfi I
GGGCCC(WGG)
GGWCC(WGG)
TGGCCA(GG)
CCWGG
YGGCCA(GG)
RGGNCCT(GG)
GGCCN5GGCC(WGG) or
GGCCWGGN2GGCC
Stu I
Van91 I
AGGCCT(GG)
CCAGGN3TGG
VpaK11B I
Enzymes NOT Blocked by Overlapping dam Methylation
Enzyme
Site
BamH I
GGATCC
Bgl II
AGATCT
Pvu I
CGATCG
Sau3A I
GATC
M = A or C; K = G or T; N = A, C, G, or T;
R = A or G; Y = C or T; W = A or T; S = G or C
GGWCCWGG
Enzymes NOT blocked by overlapping dcmmethylation
Enzyme
Site
BamH I
GGATCC
BstP I
GGTNACC
BstX I
CCAN4CCTGG
EcoO65 I
GGTNACC
Hae III
GGCC
Hin1 I
GRCGYC
Kpn I
GGTACC
Mva I
CCWGG
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