Restrictionenzymes(限制性核酸內切酶）100108128趙博BacterialenzymeswhichcutDNAintodefinedandreproduciblefragmentsIdentifiedinthelate1960sKeydiscoverywhichallowedtheDNAcloningtobecomearealityOverview限制酶—CutRestrictionenzymeorrestrictionendonuclease一種能選定特定DNA片段攻擊的核酸酶（nuclease）。許多細菌（bacteria）天生就會產生，用來自我防御。細菌產生的限制性核酸內切酶有兩種特性：只有特定的短序列（restrictionsite）會是攻擊的目標。細菌會在自己的restrictionsite(限制性內切點)加上化學保護（甲基化,methylated）。Restrictionendonuclease

(origination)Onecomponentofthebacterialrestriction-modificationsystem,anaturaldefensemechanismofbacteriatoagainsttheintroductionofforeignDNAintothecellRestrictionendonuclease:recognizeashort,symmetricalDNAsequence,andcutDNAbackboneineachstrandataspecificsitewithinthatsequence

(killforeignDNA)Methylase:methylatesCorAofthecellularDNATypesofRestrictionendonucleaseTypeITypeIITypeIIIFunctionsEndonuclease&methylaseEndonucleaseEndonucleaseConditionsATP,Mg2+Mg2+ATP,Mg2+RecognitionsequencesEcoK:AACN6GTGCEcoB:TGAN8TGCTPalindromic(回文序列）EcoP1:AGACCEcoP15:CAGCAGCuttingsitesAtleast1000bpawayAtorclosetorecog.seq24-26bpawayTypeII

Recognitionsequencecharacteristics（識別序列特點）——

palindrome（回文結構）

切口：平端切口、粘端切口GGATCCCCTAGGBamHⅠGTCCAGGCCTAGGATCCG+GGATCCCCTAGGHindⅡGTCGACCAGCTGGACCTG+Flatendincision（平端切口）Stickyendincision（粘端切口）GTCGACCAGCTG5’protrudingends3’protrudingends5’-CCCGGG-3’3’-GGGCCC-5’5’-CCC-OH3’-GGG-pp-GGG-3’OH-CCC-5’+SmaIbluntends（平末端）Cohesive/stickyends（粘性末端）Restrictionsequences（限制性識別序列）Enzymeandconnection

（酶切與連接）Commonenzymenomenclature（命名）EcoRⅠ

屬

種

株

序Escherichia

coli

RY13株大腸桿菌RY13株的第一種酶同尾酶BamHⅠBglⅡGGATCCCCTAGG

AGATCTTCTAGAGCCTAGGATCCG++ATCTAG

GATCTAisochizomer(同功異源酶)