Name Sequence Overhang Properties Isoschizomers
Aat II G↑ACGT↓C 3′ ACGT ZraI*
Acc I GT↓MK↑AC 5′ MK FblI, XmiI
Acc III T↓CCGG↑A 5′ CCGG Aor13HI, BseAI, Bsp13I, BspEI, Kpn2I, MroI
Acu I CTGAAGN₁₄↑NN↓ 3′ NN Eco57I
Afl II C↓TTAA↑G 5′ TTAA BfrI, BspTI, BstAFI, MspCI, Vha464I
Age I A↓CCGG↑T 5′ CCGG AsiGI, BshTI, CspAI, PinAI
Alw I GGATCNNNN↓N↑ 5′ N AclWI, BspPI
Alw26 I GTCTCN↓NNNN↑ 5′ NNNN BcoDI, BsmAI, BstMAI
Apa I G↓TGCA↑C 5′ TGCA Bsp120I*, PspOMI*
ApaL I G↓TGCA↑C 5′ TGCA Alw44I, VneI
Apo I R↓AATT↑Y 5′ AATT AcsI, XapI
Asc I GG↓CGCG↑CC 5′ CGCG PalAI, SgsI
Ava I C↓YCGR↑G 5′ YCGR Ama87I, BmeT110I, BsiHKCI, BsoBI, Eco88I
Ava II G↓GWC↑C 5′ GWC Bme18I, Eco47I, SinI, VpaK11BI
Avr II C↓CTAG↑G 5′ CTAG AspA2I, BlnI, XmaJI
Bal I TGG⇅CCA Blunt MlsI, MluNI, Mox20I, MscI, Msp20I
BamH I G↓GATC↑C 5′ GATC -
Bcl I T↓GATC↑A 5′ GATC FbaI, Ksp22I
Bgl I GCCN↑NNN↓NGGC 3′ NNN -
Bgl II A↓GATC↑T 5′ GATC -
Bsa I GGTCTCN↓NNNN↑ 5′ NNNN Bso31I, BspTNI, Eco31I
BsaW I W↓CCGG↑W 5′ CCGG -
BsiW I C↓GTAC↑G 5′ GTAC Pfl23II, PspLI
BsmB I CGTCTCN↓NNNN↑ 5′ NNNN Esp3I
BsoB I C↓YCGR↑G 5′ YCGR Ama87I, AvaI, BmeT110I, BsiHKCI, Eco88I
BspE I T↓CCGG↑A 5′ CCGG AccIII, Aor13HI, BseAI, Bsp13I, Kpn2I, MroI
BsrF I R↓CCGG↑Y 5′ CCGG Bse118I, BssAI, Cfr10I
BstY I R↓GATC↑Y 5′ GATC BstX2I, MflI, PsuI
BtsC I GGATG↑NN↓ 3′ NN BseGI, BstF5I, FokI*
Cfr10 I R↓CCGG↑Y 5′ CCGG Bse118I, BsrFI, BssAI
Cfr42 I CC↑GC↓GG 3′ GC KspI, SacII, Sfr303I, SgrBI
Cfr9 I C↓CCGG↑G 5′ CCGG SmaI*, TspMI, XmaI
Cla I AT↓CG↑AT 5′ CG Bsa29I, BseCI, BshVI, BspDI, Bsu15I, BsuTUI
CviA I ↓GATC↑ 5′ GATC -
Dde I C↓TNA↑G 5′ TNA BstDEI, HpyF3I
Dpn I GA⇅TC Blunt MalI
Dpn II ↓GATC↑ 5′ GATC Bsp143I, BssMI, BstKTI*, BstMBI, Kzo9I, MboI, NdeII, Sau3AI
Dra I TTT⇅AAA Blunt -
Eag I C↓GGCC↑G 5′ GGCC BseX3I, BstZI, EclXI, Eco52I
Eco47 I G↓GWC↑C 5′ GWC AvaII, Bme18I, SinI, VpaK11BI
EcoN I CCTNN↓N↑NNAGG 5′ N BstENI, XagI
EcoO109 I RG↓GNC↑CY 5′ GNC -
EcoR I G↓AATT↑C 5′ AATT -
EcoR V GAT⇅ATC Blunt Eco32I
EcoT38 I G↑RGCY↓C 3′ RGCY BanII, Eco24I, FriOI
Esp3 I CGTCTCN↓NNNN↑ 5′ NNNN BsmBI
Fok I GGATGN₉↓NNNN↑ 5′ NNNN BseGI*, BstF5I*, BtsCI*
Fsp I TGC⇅GCA Blunt Acc16I, NsbI
Hae II R↑GCGC↓Y 3′ GCGC BfoI, BstH2I
Hae III GG⇅CC Blunt BshFI, BsnI, BspANI, BsuRI
Hga I GACGCN₅↓NNNNN↑ 5′ NNNNN CseI
Hinc II GTY⇅RAC Blunt HindII
XmaI C↓CCGG↑G 5′ CCGG Cfr9I, SmaI*, TspMI
Hind II GTY⇅RAC Blunt HincII
Hind III A↓AGCT↑T 5′ AGCT -
Hinf I G↓ANT↑C 5′ ANT -
HinP1 I G↓CG↑C 5′ CG AspLEI*, BstHHI*, CfoI*, HhaI*, Hin6I, HspAI
Hpa I GTT⇅AAC Blunt KspAI
Hpa II C↓CG↑G 5′ CG BsiSI, HapII, MspI
Hph I GGTGAN₇↑N↓ 3′ N AsuHPI
Hpy188 I TC↑N↓GA 3′ N -
Hpy99 I ↑CGWCG↓ 3′ CGWCG -
HpyCH4 V TG⇅CA Blunt HpySE526I, MaeII, TaiI*
Kpn I G↑GTAC↓C 3′ GTAC Acc65I*, Asp718I*
Kpn2 I T↓CCGG↑A 5′ CCGG AccIII, Aor13HI, BseAI, Bsp13I, BspEI, MroI
Lsp1109 I GCAGCN₈↓NNNN↑ 5′ NNNN BbvI, BseXI, BstV1I
Mbo I ↓GATC↑ 5′ GATC Bsp143I, BssMI, BstKTI*, BstMBI, DpnII, Kzo9I, NdeII, Sau3AI
Mbo II GAAGAN₇↑N↓ 3′ N -
Mlu I A↓CGCG↑T 5′ CGCG -
Xho I C↓TCGA↑G 5′ TCGA PaeR7I, Sfr274I, SlaI
Mnl I CCTCN₆↑N↓ 3′ N -
Mse I T↓TA↑A 5′ TA SaqAI, Tru1I, Tru9I
Msp I C↓CG↑G 5′ CG BsiSI, HapII, HpaII
MspA1 I CMG⇅CKG Blunt -
Mun I C↓AATT↑G 5′ AATT MfeI
Nae I GCC⇅GGC Blunt MroNI*, NgoMIV*, PdiI
Nco I C↓CATG↑G 5′ CATG Bsp19I
Nde I CA↓TA↑TG 5′ TA FauNDI
NgoM IV G↓CCGG↑C 5′ CCGG MroNI, NaeI*, PdiI*
Nhe I G↓CTAG↑C 5′ CTAG AsuNHI, BmtI*, BspOI*
Nla IV GGN⇅NCC Blunt BmiI, BspLI, PspN4I
Not I GC↓GGCC↑GC 5′ GGCC CciNI
Nru I TCG⇅CGA Blunt Bsp68I, BtuMI, RruI
Nt.BstNB I GAGTCNNNN↓ Not applicable -
PaeR7 I C↓TCGA↑G 5′ TCGA Sfr274I, SlaI, XhoI
PflM I CCAN↑NNN↓NTGG 3′ NNN AccB7I, Van91I
Ple I GAGTCNNNN↓N↑ 5′ N MlyI*, PpsI, SchI*
PluT I G↑GCGC↓C 3′ GCGC DinI*, EgeI*, EheI*, KasI, SfoI*
PspG I ↓CCWGG↑ 5′ CCWGG AjnI, BciT130I*, BseBI*, BstNI*, Bst2UI*, EcoRII, MvaI*, Psp6I
Xba I T↓CTAG↑A 5′ CTAG -
Tth111 I GACN↓N↑NGTC 5′ N PflFI, PsyI
TspM I C↓CCGG↑G 5′ CCGG Cfr9I, SmaI*, XmaI
Taq I T↓CG↑A 5′ CG -
Swa I ATTT⇅AAAT Blunt SmiI
Pst I C↑TGCA↓G 3′ TGCA BspMAI
Pvu I CG↑AT↓CG 3′ AT Ple19I
Pvu II CAG⇅CTG Blunt -
Rsa I GT⇅AC Blunt AfaI, Csp6I*, CviQI*, RsaNI*
Sac I G↑AGCT↓C 3′ ACGT Psp124BI, SstI, Ecl136II*, EcoICRI*, Eco53kI*
Sac II CC↑GC↓GG 3′ GC Cfr42I, KspI, Sfr303I, SgrBI
Sal I G↓TCGA↑C 5′ TCGA -
Sau96 I G↓GNC↑C 5′ GNC AspS9I, BmgT120I, Cfr13I, PspPI
Sbf I CC↑TGCA↓GG 3′ TGCA SdaI, Sse8387I
Sca I AGT⇅ACT Blunt ZrmI
Sda I CC↑TGCA↓GG 3′ TGCA SbfI, Sse8387I
Sfi I GGCCN↑NNN↓NGGCC 3′ NNN -
SgrA I CR↓CCGG↑YG 5′ CCGG -
Sma I CCC⇅GGG Blunt Cfr9I*, TspMI*, XmaI*
SnaB I TAC⇅GTA Blunt BstSNI, Eco105I
Spe I A↓CTAG↑T 5′ CTAG AhlI, BcuI
Sph I G↑CATG↓C 3′ CATG PaeI
Sse9 I ↓AATT↑ 5′ AATT MluCI, TasI
Ssp I AAT⇅ATT Blunt -
Stu I AGG⇅CCT Blunt Eco147I, PceI, SseBI
StyD4 I ↓CCNGG↑ 5′ CCNGG Bme1390I*, BmrFI*, BstSCI, MspR9I*, ScrFI*

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.

 

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.