Enzyme-Finder

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Ambiguity Codes

n
A or C or G or T (any)
b
C or G or T (not A)
d
A or G or T (not C)
h
A or C or T (not G)
v
A or C or G (not T)
w
A or T (weak)
s
C or G (strong)
r
A or G (purine)
y
C or T (pyrimidine)
m
A or C (amino)
k
G or T (keto)

Name	Sequence ?	Overhang	Properties ?	Isoschizomere
Aat II	G↑ACGT↓C	3′ ACGT	37° IV 65° CpG FcB	ZraI*
Acc I	GT↓MK↑AC	5′ MK	37° IV 80° CpG FcB	Bsh1236I, BspFNI, BstFNI, BstUI, MvnI
Acc III	T↓CCGG↑A	5′ CCGG	65° AccIII No Dam	Aor13HI, BseAI, Bsp13I, BspEI, Kpn2I, MroI
Acu I	CTGAAGN₁₄↑NN↓	3′ NN	37° IV 65° FcB	Eco57I
Afl II	C↓TTAA↑G	5′ TTAA	37° IV 65° FcB	BfrI, BspTI, BstAFI, MspCI, Vha464I
Age I	A↓CCGG↑T	5′ CCGG	37° IV 65° CpG FcB	AsiGI, BshTI, CspAI, PinAI
Alw I	GGATCNNNN↓N↑	5′ N	37° IV No Dam FcB	AclWI, BspPI
Alw26 I	GTCTCN↓NNNN↑	5′ NNNN	37° IV 65° CpG FcB	BcoDI, BsmAI, BstMAI
Apa I	G↑GGCC↓C	3′ GGCC	25° IV 65° CpG Dcm FcB	Bsp120I, PspOMI
ApaL I	G↓TGCA↑C	5′ TGCA	37° IV No CpG FcB	Alw44I, VneI
Apo I	R↓AATT↑Y	5′ AATT	50° III 80° FcB	AcsI, XapI
Asc I	GG↓CGCG↑CC	5′ CGCG	37° IV 65° CpG FcB	PalAI, SgsI
Ava I	C↓YCGR↑G	5′ YCGR	37° IV 80° CpG FcB	Ama87I, BmeT110I, BsiHKCI, BsoBI, Eco88I
Ava II	G↓GWC↑C	5′ GWC	37° IV 80° CpG Dcm FcB	Bme18I, Eco47I, SinI, VpaK11BI
Avr II	C↓CTAG↑G	5′ CTAG	37° IV 80° FcB	AspA2I, BlnI, XmaJI
Bal I	TGG⇅CCA	Blunt	37° BalI 65° Dcm	MlsI, MluNI, Mox20I, MscI, Msp20I
BamH I	G↓GATC↑C	5′ GATC	37° BamHI No FcB	-
Bcl I	T↓GATC↑A	5′ GATC	50° III No Dam FcB	FbaI, Ksp22I
Bgl I	GCCN↑NNN↓NGGC	3′ NNN	37° III 65° CpG FcB	-
Bgl II	A↓GATC↑T	5′ GATC	37° III No FcB	-
Bsa I	GGTCTCN↓NNNN↑	5′ NNNN	37° IV 65° CpG Dcm FcB	Bso31I, BspTNI, Eco31I
BsaW I	W↓CCGG↑W	5′ CCGG	60° IV 80° FcB	-
BsiW I	C↓GTAC↑G	5′ GTAC	55° III 80° CpG FcB	Pfl23II, PspLI
BsmB I	CGTCTCN↓NNNN↑	5′ NNNN	55° III 80° CpG FcB	Esp3I
BsoB I	C↓YCGR↑G	5′ YCGR	37° IV 80° FcB	Ama87I, AvaI, BmeT110I, BsiHKCI, Eco88I
BspE I	T↓CCGG↑A	5′ CCGG	37° III 80° CpG Dam FcB	AccIII, Aor13HI, BseAI, Bsp13I, Kpn2I, MroI
BsrF I	R↓CCGG↑Y	5′ CCGG	37° IV No CpG FcB	Bse118I, BssAI, Cfr10I
BstY I	R↓GATC↑Y	5′ GATC	60° II 80° FcB	BstX2I, MflI, PsuI
BtsC I	GGATG↑NN↓	3′ NN	50° IV 80° FcB	BseGI, BstF5I, FokI*
Cfr10 I	R↓CCGG↑Y	5′ CCGG	37° Cfr10I No CpG FcB	Bse118I, BsrFI, BssAI
Cfr42 I	CC↑GC↓GG	3′ GC	37° I 65° CpG FcB	KspI, SacII, Sfr303I, SgrBI
Cfr9 I	C↓CCGG↑G	5′ CCGG	37° III 65° CpG	SmaI*, TspMI, XmaI
Cla I	AT↓CG↑AT	5′ CG	37° IV 65° CpG Dam FcB	Bsa29I, BseCI, BshVI, BspDI, Bsu15I, BsuTUI
CviA I	↓GATC↑	5′ GATC	37° IV 65° Dam FcB	-
Dde I	C↓TNA↑G	5′ TNA	37° III 65° FcB	BstDEI, HpyF3I
Dpn I	GA⇅TC	Blunt	37° IV 80° FcB	MalI
Dpn II	↓GATC↑	5′ GATC	37° DpnII 65° Dam FcB	Bsp143I, BssMI, BstKTI*, BstMBI, Kzo9I, MboI, NdeII, Sau3AI
Dra I	TTT⇅AAA	Blunt	37° IV 65° FcB	-
Eag I	C↓GGCC↑G	5′ GGCC	37° III 65° CpG FcB	BseX3I, BstZI, EclXI, Eco52I
Eco47 I	G↓GWC↑C	5′ GWC	37° III 65° CpG Dcm FcB	AvaII, Bme18I, SinI, VpaK11BI
EcoN I	CCTNN↓N↑NNAGG	5′ N	37° IV 65° FcB	BstENI, XagI
EcoO109 I	RG↓GNC↑CY	5′ GNC	37° IV 65° Dcm FcB	-
EcoR I	G↓AATT↑C	5′ AATT	37° EcoRI 65° CpG FcB	-
EcoR V	GAT⇅ATC	Blunt	37° III 65° CpG FcB	Eco32I
EcoT38 I	G↑RGCY↓C	3′ RGCY	37° IV 65° FcB	BanII, Eco24I, FriOI
Esp3 I	CGTCTCN↓NNNN↑	5′ NNNN	37° IV 65° CpG FcB	BsmBI
Fok I	GGATGN₉↓NNNN↑	5′ NNNN	37° IV 65° CpG Dcm FcB	BseGI, BstF5I, BtsCI*
Fsp I	TGC⇅GCA	Blunt	37° IV 65° CpG FcB	Acc16I, NsbI
Hae II	R↑GCGC↓Y	3′ GCGC	37° IV 80° CpG FcB	BfoI, BstH2I
Hae III	GG⇅CC	Blunt	37° IV 80° FcB	BshFI, BsnI, BimgI, BsuRI
Hga I	GACGCN₅↓NNNNN↑	5′ NNNNN	37° I 65° CpG FcB	CseI
Hinc II	GTY⇅RAC	Blunt	37° IV 65° CpG FcB	HindII
XmaI	C↓CCGG↑G	5′ CCGG	37° IV 65° CpG FcB	Cfr9I, SmaI*, TspMI
Hind II	GTY⇅RAC	Blunt	37° II 65° CpG FcB	HincII
Hind III	A↓AGCT↑T	5′ AGCT	37° II 80° FcB	-
Hinf I	G↓ANT↑C	5′ ANT	37° IV 80° CpG FcB	-
HinP1 I	G↓CG↑C	5′ CG	37° II 65° CpG FcB	AspLEI, BstHHI, CfoI, HhaI, Hin6I, HspAI
Hpa I	GTT⇅AAC	Blunt	37° IV No CpG FcB	KspAI
Hpa II	C↓CG↑G	5′ CG	37° IV 80° CpG FcB	BsiSI, HapII, MspI
Hph I	GGTGAN₇↑N↓	3′ N	37° IV 65° Dam FcB	AsuHPI
Hpy188 I	TC↑N↓GA	3′ N	37° IV 65° Dam FcB	-
Hpy99 I	↑CGWCG↓	3′ CGWCG	37° IV 65° CpG FcB	-
HpyCH4 V	TG⇅CA	Blunt	37° IV 65° FcB	-
Kpn I	G↑GTAC↓C	3′ GTAC	37° I No FcB	Acc65I, Asp718I
Kpn2 I	T↓CCGG↑A	5′ CCGG	55° I 80° CpG FcB	AccIII, Aor13HI, BseAI, Bsp13I, BspEI, MroI
Lsp1109 I	GCAGCN₈↓NNNN↑	5′ NNNN	37° III 65° FcB	BbvI, BseXI, BstV1I
Mbo I	↓GATC↑	5′ GATC	37° III 65° CpG Dam FcB	Bsp143I, BssMI, BstKTI*, BstMBI, DpnII, Kzo9I, NdeII, Sau3AI
Mbo II	GAAGAN₇↑N↓	3′ N	37° II 65° Dam FcB	-
Mlu I	A↓CGCG↑T	5′ CGCG	37° III 65° CpG FcB	-
Xho I	C↓TCGA↑G	5′ TCGA	37° IV 80° CpG FcB	PaeR7I, Sfr274I, SlaI
Mnl I	CCTCN₆↑N↓	3′ N	37° II 65° FcB	-
Mse I	T↓TA↑A	5′ TA	37° IV 65° FcB	SaqAI, Tru1I, Tru9I
Msp I	C↓CG↑G	5′ CG	37° IV No FcB	BsiSI, HapII, HpaII
MspA1 I	CMG⇅CKG	Blunt	37° IV 65° CpG FcB	-
Mun I	C↓AATT↑G	5′ AATT	37° II 65° FcB	MfeI
Nae I	GCC⇅GGC	Blunt	37° I 65° CpG FcB	MroNI, NgoMIV, PdiI
Nco I	C↓CATG↑G	5′ CATG	37° III 65° FcB	Bsp19I
Nde I	CA↓TA↑TG	5′ TA	37° IV 65° FcB	FauNDI
NgoM IV	G↓CCGG↑C	5′ CCGG	37° IV 80° CpG FcB	MroNI, NaeI, PdiI
Nhe I	G↓CTAG↑C	5′ CTAG	37° II 65° CpG FcB	AsuNHI, BmtI, BspOI
Nla IV	GGN⇅NCC	Blunt	37° IV 65° CpG Dcm FcB	BmiI, BspLI, PspN4I
Not I	GC↓GGCC↑GC	5′ GGCC	37° III 65° CpG FcB	CciNI
Nru I	TCG⇅CGA	Blunt	37° III 65° CpG Dam FcB	Bsp68I, BtuMI, RruI
Nt.BstNB I	GAGTCNNNN↓	Nicht vorhanden	55° III 80° FcB	-
PaeR7 I	C↓TCGA↑G	5′ TCGA	37° IV No CpG FcB	Sfr274I, SlaI, XhoI
PflM I	CCAN↑NNN↓NTGG	3′ NNN	37° III 65° Dcm FcB	AccB7I, Van91I
Ple I	GAGTCNNNN↓N↑	5′ N	37° IV 65° CpG FcB	MlyI, PpsI, SchI
PluT I	G↑GCGC↓C	3′ GCGC	37° IV 65° CpG FcB	DinI, EgeI, EheI, KasI, SfoI
PspG I	↓CCWGG↑	5′ CCWGG	37° IV No Dcm	AjnI, BciT130I, BseBI, BstNI, Bst2UI, EcoRII, MvaI*, Psp6I
Pst I	C↑TGCA↓G	3′ TGCA	37° III 80° FcB	BspMAI
Xba I	T↓CTAG↑A	5′ CTAG	37° IV 65° Dam FcB	-
Tth111 I	GACN↓N↑NGTC	5′ N	65° IV No FcB	PflFI, PsyI
TspM I	C↓CCGG↑G	5′ CCGG	75° IV No CpG FcB	Cfr9I, SmaI*, XmaI
Taq I	T↓CG↑A	5′ CG	65° III 80° Dam FcB	-
Swa I	ATTT⇅AAAT	Blunt	25° III 65° FcB	SmiI
Pvu I	CG↑AT↓CG	3′ AT	37° III No CpG FcB	Ple19I
Pvu II	CAG⇅CTG	Blunt	37° II No FcB	-
Rsa I	GT⇅AC	Blunt	37° IV 65° CpG FcB	AfaI, Csp6I, CviQI, RsaNI*
Sac I	G↑AGCT↓C	3′ AGCT	37° I 65° FcB	Ecl136II, EcoICRI, Eco53kI*, Psp124BI, SstI
Sac II	CC↑GC↓GG	3′ GC	37° IV 65° CpG FcB	Cfr42I, KspI, Sfr303I, SgrBI
Sal I	G↓TCGA↑C	5′ TCGA	37° III 65° CpG FcB	-
Sau96 I	G↓GNC↑C	5′ GNC	37° IV 80° CpG Dcm FcB	AspS9I, BmgT120I, Cfr13I, PspPI
Sbf I	CC↑TGCA↓GG	3′ TGCA	37° IV 80° FcB	SdaI, Sse8387I
Sca I	AGT⇅ACT	Blunt	37° III 80° FcB	ZrmI
Sda I	CC↑TGCA↓GG	3′ TGCA	37° IV 80° FcB	SbfI, Sse8387I
Sfi I	GGCCN↑NNN↓NGGCC	3′ NNN	50° II No CpG Dcm FcB	-
SgrA I	CR↓CCGG↑YG	5′ CCGG	37° IV 65° CpG FcB	-
Sma I	CCC⇅GGG	Blunt	25° IV 65° CpG FcB	Cfr9I, TspMI, XmaI*
SnaB I	TAC⇅GTA	Blunt	37° IV 80° CpG FcB	BstSNI, Eco105I
Spe I	A↓CTAG↑T	5′ CTAG	37° IV 80° FcB	AhlI, BcuI
Sph I	G↑CATG↓C	3′ CATG	37° II 65° FcB	PaeI
Sse9 I	↓AATT↑	5′ AATT	55° I 65° FcB	MluCI, TasI
Ssp I	AAT⇅ATT	Blunt	37° IV 65° FcB	-
Stu I	AGG⇅CCT	Blunt	37° IV 65° Dcm FcB	Eco147I, PceI, SseBI
StyD4 I	↓CCNGG↑	5′ CCNGG	37° IV 65° CpG Dcm FcB	Bme1390I, BmrFI, BstSCI, MspR9I, ScrFI

Double digest − Buffer compatibility chart

Performing two digestions simultaneously saves a lot of time.

Check the buffer compatibility at the FastGene^® Double Digest Chart:

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Restriction Endonucleases

What are Restriction Enzymes?

Restriction enzmyes recognize short DNA sequences and cleaves double-stranded DNA at or near a specific recognition site. Restriction enzmyes are classified into four types, based on their subunit structure, cofactor requirements and specificity of cleavage.

3,000 different restriction enzymes have been discovered, which recognize over 230 distinct DNA sequences. These enzymes are routinely used for DNA modification around the world and are an indispensable tool in molecular cloning.

Historical background

The basis for the research of restriction enzymes goes back to the work of Luria and colleagues in the early 1950s [1]. Luria observed that the bacteriophage λ can grow good in one strain of E.coli (e.g. E.coli C), but often poorly in another E.coli strain (e.g. E.coli K). The host cell (E.coli K) was known as the restriction host and appears to have the ability to reduce the biological activity of the phage λ.

The first-time that the term restriction enzyme was mentioned was in the 1960s in the laboratories of Arber and Meselson. They found out that the restriction is caused by an enzymatic cleavage of the phage DNA. The enzyme involved in this process was termed “restriction enzyme” [2, 3]. The restriction enzymes studied by Arber and Meselson were type I restriction enzymes, which cleave DNA at random places away from the recognition site.

In 1970, Smith and colleagues isolated and describe the first type II restriction enzyme, Hind II [4]. Restriction enzymes of type II are much more useful for laboratory work, because they cleave DNA at the site of their recognition sequence. Due to its importance for molecular biology, Smith, Arber and Nathans shared the 1978 Nobel Prize for Medicine and Physiology for their discovery of restriction enzymes and their application to molecular genetics.

Recognition sequences

All restriction endonucleases recognize a specific DNA sequence. The recognition sequence is usually palindromic or partially palindromic, meaning the base sequence reads the same backwards and forwards. Restriction enzymes can cleave double stranded DNA either at the center of both strands to yield “blunt ends” or at a staggered position leaving overhangs called “sticky ends”.

Different Types of restriction enzymes

Based on the structure, cofactor requirements and specificity of cleavage there are four types of restriction enzymes (Types I, II, III, and IV).

Type I restriction enzymes cleaves the DNA at a random location far away from the recognition sequence. These enzymes require both ATP and S-adenosyl-L-methionine to function.

Type II restriction enzymes cleaves the DNA within or near the recognition sequence. These enzymes do not require ATP and are independent from methylase. The type II enzymes are the most useful restriction endonucleases for the daily laboratory work. All our NIPPON Genetics EUROPE restriction enzymes are from type II.

Type III restriction enzymes cleaves DNA about 20 – 25 base pairs away from the recognition sequence. They require both ATP and S-adenosyl-L-methionine to function.

Type IV restriction enzymes cleaves only modified, typically methylated DNA in contrast to the types I-III, which are usually inhibited by methylation.

Double digestion

A vector and an insert DNA can be cloned by cleaving with two different restriction enzymes, thus generating two different restriction ends. This strategy prevents the vector from being ligated without an insert, resulting in great reduction in self-ligation and increase in cloning efficiency. Most of our restriction enzymes are 100% active in the FastCut Buffer, making double digestion simple. Please look at the Double Digest Chart to make a double digestion with the four standard buffers.

DOUBLE DIGEST CHART (DOWNLOAD)

Activity of Unique FastGene^® Buffer

Nippon Genetics provides four standard buffers that maximally support the activity of each restriction enzyme in the buffer provided with the enzyme. However, some restriction endonucleases require unique buffer for maximal activity. Take a look on the Double Digest Chart to select a buffer for double digestion if a restriction enzyme requires unique buffer. Listed are activities (%) of the most commonly used restriction endonucleases in the five unique FastGene^® Buffers for EcoR I, BamH l, Acc III, Bal I, and Dpn Il, respectively. A restriction enzyme, if it is active in one of the four standard buffers, is usually active in a unique buffer. Therefore, it is possible to perform double digestion in a particular unique buffer. If digestion efficiency is low due to the suboptimal buffer, increase the amount of restriction endonucleases or incubate for a longer period of time.

DOUBLE DIGEST CHART (DOWNLOAD)

References

[1] Luria and Human (1952) A nonhereditary, host-induced variation of bacterial viruses. J Bacteriol., 557-569.

[2] Arber and Linn (1969) DNA modification and restriction. Annu Rev Biochem., 467-500.

[3] Meselson and Yuan (1968) DNA restriction enzyme from E. coli. Nature, 1110-1114

[4] Smith and Wilcox (1970) A restriction enzyme from Hemophilus influenzae. I. Purification and general properties. J Mol Biol., 379-391.