请使用支持JavaScript的浏览器!
主营:连接蛋白、DNA修饰酶、NGS文库制备(NGS Library Construction)、核酸和RNA酶。
banner
当前位置: 首页 > 产品中心 > Ultrapure Ligases > Enzymatics/T4 DNA Ligase (Rapid)/L6030-HC-L/240,000 U
公司介绍
Enzymatics is an emerging leader in the supply of the enzymes that drive nucleic acid detection technologies. Our leadership is currently building a team which embodies our mission to lead the molecular biology tools market in quality, customer servi...

Enzymatics/T4 DNA Ligase (Rapid)/L6030-HC-L/240,000 U

Enzymatics/T4 DNA Ligase (Rapid)/L6030-HC-L/240,000 U
  • Enzymatics/T4 DNA Ligase (Rapid)/L6030-HC-L/240,000 U
商品介绍

T4DNALigase(Rapid)

ProductDescription

T4DNALigase catalyzestheformationofaphosphodiesterbondbetweentheterminal5′phosphateanda3′hydroxylgroupsofduplexDNAorRNA.TheenzymeefficientlyjoinsbluntandcohesiveendsandrepairssinglestrandednicksinduplexDNA,RNAorDNA/RNAhybrids(1).

SourceofProtein
Arecombinant E.coli straincarryingtheclonedT4DNALigasegene.

SuppliedIn
10mMTris-HCl
50mMKCl
1mMDTT
0.1mMEDTA
50%glycerol
pH7.5@25°C

SuppliedWith
B1010(2XRapidLigationBuffer)
B6030(10XT4DNALigaseBuffer)

2XRapidLigationBuffer(B1010):
132mMTris-HCI
20mMMgCl2
2mMDTT
2mMATP
15%PEG6000
pH7.6@25°C

10XT4DNALigaseBuffer(B6030):
500mMTris-HCI
100mMMgCl2
50mMDTT
10mMATP
pH7.6@25°C

UnitDefinition
1unitisdefinedastheamountofDNALigaserequiredtojoin50%of100ngofDNAfragmentswithcohesiveterminiin50µl1XDNALigaseBufferfollowinga30minuteincubationat23°C.

QualityControlAnalysis

UnitCharacterizationAssay
Unitactivitywasmeasuredusinga2-foldserialdilutionmethod.Dilutionsofenzymeweremadein1XDNALigaseReactionBufferandaddedto20µLreactionscontainingdoublestrandedDNAfragmentsand1XDNALigaseReactionBuffer.Reactionsareincubatedfor30minutesat~23°C,stopped,andanalyzedona1%agarosegelstainedwithethidiumbromide.

ProteinConcentration(OD280)Measurement
A2.0µLsampleofenzymewasanalyzedatOD280 usingaNanodropND-1000spectrophotometerstandardizedusinga2.0mg/mlBSAsample(PierceCat#23209)andblankedwithproductstoragesolution.Theobservedaveragemeasurementof3replicatesampleswasconvertedtomg/mLusinganextinctioncoefficientof54,050andmolecularweightof55,292Daltons.

SDS-Page(PhysicalPurityAssessment)
2.0µLofconcentratedenzymesolutionwasloadedonadenaturing4-20%Tris-GlycineSDS-PAGEgelflankedbyabroad-rangeMWMarkerand2.0µLofa1:100dilutionofthesameenzymespecies.Followingelectrophoresis,thegelwasstainedandthesamplescomparedtodeterminephysicalpurity.Theacceptancecriteriaforthistestrequiresthattheaggregatemassofcontaminantbandsintheconcentratedsampledonotexceedthemassoftheproteinofinterestbandinthedilutesample,confirminggreaterthan99%purityoftheconcentratedsample.

ContaminationTests

Single-StrandedExonucleaseActivity
A50µLreactioncontaining10,000cpmofarADIolabeledsingle-strandedDNAsubstrateand10µLofenzymesolutionincubatedfor4hoursat37°Cresultedinlessthan1.0%releaseofTCA-solublecounts.

Double-StrandedExonucleaseActivity
A50µlreactioncontaining5,000cpmofaradiolabeleddouble-strandedDNAsubstrateand10µLofenzymesolutionincubatedfor4hoursat37°Cresultedinlessthan1.0%releaseofTCA-solublecounts.

Double-StrandedEndonucleaseActivity
A50µLreactioncontaining0.5µgofpENZuCDNAand10µLofenzymesolutionincubatedfor4hoursat37°CresultedinnovisuallydiscernIBLeconversiontonickedcircularDNAasdeterminedbyagarosegelelectrophoresis.

E.coli 16SrDNAContaminationTest:
Replicate5µLsamplesofenzymesolutionweredenaturedandscreenedinaTaqManqPCRassayforthepresenceofcontaminating E.coli genomicDNAusingoligonucleotideprimerscorrespondingtothe16SrRNAlocus.Theacceptancecriterionforthetestisthethresholdcyclecount(Ct)producedbytheaverageof3replicatenotemplatecontrolsamples.BasedonthecorrelationbetweenthenotemplatecontrolCt valuesandstandardcurvedata,thedetectionlimitofthisassayis<10copiesgenome/sample.

ProductInformation

T4DNALigase(Rapid)
PartNumberL6030-HC-L
Price$513
Concentration600,000U/ml
UnitSize240,000U
SDSAvailableonrequest

ProductSpecification*

StorageTemperature-25to-15°C
TestSpecification
Purity(SDS-PAGE)>99%
SpecificActivity300,000U/mg
SSExonuclease6,000U<1.0%released
DSExonuclease6,000U<1.0%released
DSEndonuclease6,000U=noconversion
E.coliDNAContamination6,000U<10copies

*Foradetailedsummaryofassayconditionsanddata,refertotheQualityControlsAnalysissection.

UsageInstructions

ReactionSet-Up:

AmountDescriptionFinalConcentration
10µL2XRapidLigationBuffer1X
XµLVector1-10ng/µL
XµLInsert1-10ng/µL
1µLT4DNALigase(600U/µL)30U/µL
XµLTypeIWaterN/A
20µLTotalVolume

 

  1. Addallofabovecomponentstoacleanreactionvessel,mixwellbypipetting.
  2. Incubateat25°Cfor10minutes.
  3. ImmediatelypurifyDNAusingPCRclean-upcolumnandelutein~50µL.
  4. –OR–Immediatelydilute(atleast1:10,butenoughsuchthat0.1-10ngofligationproductwillbetransformed)inTEorwater
  5. Transform0.1-10ngofligationproductintochemicallyorelectrocompetentcelllinethatiscompatiblewithvector

Notes

OneEnzymaticsT4DNALigasecohesiveendunitisequivalenttoapproximately3cohesiveendunitsasmeasuredwithaLamBDa-HindIIIDNAfragmentsubstratein1XT4DNALigasereactionbuffer.

OneWeissUnitisapproximatelyequivalentto22Enzymaticscohesiveendunits.

T4DNALigaseisATPdependent.Itisrecommendedthatthereactionbufferbediscardedafteroneyearofstorageat-20°Candreplacedwithfreshbuffertoensuremaximumperformance.Single-insertligationsareoptimalwhentargetinganinsert:vectorratiobetween2and6.Aratioabove6:1willpromotetheinsertionofmultiplefragments,whiledroppingbelow2:1willreduceligationefficiency.

ForproblematicligationsoriftheDNAconcentrationisunknown,itmaybenecessarytovaryratiosandrunmultipleligations.

ThepresenceofPEGatahighconcentrationwillsignificantlyreducethetransformationefficiencyofelectrocompetentcells.Inordertomaximizetheefficiencyoftransformationintoelectrocompetentcells,thefollowingapproachesarerecommended:

Best:Followingligation,purifytheproductusingaDNApurificationspincolumnandelutein50µLofTE.TheDNAisnowreadyfortransformation.ThefinalamountofDNAtobetransformedshouldbeintherangeof0.1-10ng.

Better:DiluteligationproductinddH20orTEtoreducethePEGconcentration.ThefinalamountofDNAtobetransformedshouldbeintherangeof0.1-10ng.

Enzymatics’high-concentrationT4DNALigaseincombinationwiththe2XRapidLigationbuffergreatlystimulatestherateandefficiencyblunt-endligation,thereforelongincubations(>10minutes)areNOTrecommendedandcangreatlyreducethetransformationefficiencyofligationproducts.Inordertomaximizetransformationefficiencyofthecorrectinsert/vectorcombination,thefollowingprotocolisrecommended.

Enzymatics10XT4DNALigaseBufferdoesnotcontainPEGandiscompatiblewithstandardligationprotocolswhichdonotspecifytheuseofarapid/fast/quickformatbuffer.

References

  1. Engler,M.J.andRichardson,C.C.(1982)P.D.Boyer(Eds.),TheEnzymes,5,pp.3.SanDiego:AcademicPress.

LimitationsofUse
Thisproductwasdeveloped,manufactured,andsoldforinvitrouseonly.TheproductisnotsuitableforadmiNISTrationtohumansoranimals.SDSsheetsrelevanttothisproductareavailableuponrequest.

品牌介绍

Enzymatics is an emerging leader in the supply of the enzymes that drive nucleic acid detection technologies. Our leadership is currently building a team which embodies our mission to lead the molecular biology tools market in quality, customer service, and innovation. We seek engaging, motivated individuals who can articulate their goals and, in turn, offer a culture that breeds success. Enzymatics is an employee-focused organization that strives to cultivate challenging careers and rewarding lives. If you seek a challenge, have a thirst for knowledge, and are interested in having fun while working, please apply to learn more!

We offer a competitive benefits package and an incentive-based compensation package.



苏州蚂蚁淘生物科技有限公司是一家创新型高科技生物公司,以常规生物学实验外包与生物学试剂为基础,放眼于生命科学领域的前沿技术服务,公司技术骨干有着多年的生物学科研背景,毕业于国内外名牌大学,并且有着丰富的项目经验,了解国内外科研行情,我公司力求为每一个客户提专业、针对性的实验方案。我公司以技术服务为支点,和国内外多家生物试剂公司有着密切的业务往来.公司特色技术服务包括:抗体药物研发、噬菌体抗体库、蛋白表达与传化、抗体制备、高通量测序、组学研究、生物信息学分析与方法建立、基础分子生物学技术外包、化学中间体合成、原料药物研发、论文编写等。我公司秉承”诚信做人,认真做事,服务至上“,希望用我们的专业知识和项目经验为国内科研客户解决科研问题,祝您科研之路畅通无阻!


Concentration 200,000U/mL

UnitSize 10,000U

LotNumber Shipmentdependent

StorageTemperature-25°Cto-15°C

SDSPurity n/a >99%

SpecificActivity n/a ≥280,000U/mg

E.coliDNAContamination 2,000 <10copies

核酸内切酶

摘要:能够降解RNA/DNA杂交链中的RNA链。

大肠杆菌RNaseH(rnh)属于核酸内切酶,该酶能够降解RNA/DNA杂交链中的RNA链。该酶的降解产物核苷酸分子具有5-磷酸末端和3-羟基末端。该酶对于单链或双链RNA分子几乎无活性

产品组分

SuppliedWith

B9220(10XRNAseHBuffer)

10XRNAseHBuffer(B9220):

500mMTris-HCl

750mMKCl

30mMMgCl2

100mMDTT

pH8.3@25°C

产品信息:

PartNumber Y9220L,Y9220F

Concentration 5,000U/ml

UnitSize 5,000U,770U

StorageTemperature -25to-15°C

Purity(SDS-PAGE) >99%

SpecificActivity 625,000U/mg

E.coliDNAContamination 500U<10copies

T4DNA聚合酶

摘要:为DNA克隆期间抛光5′和3′端提供可靠选择

T4DNA聚合酶能催化原始DNA模板以5′→3′方向延伸。该酶具有较强的3′→5′核酸外切酶活性,但缺乏内在的5′→3′核酸外切酶活性或链置换功能。

Suppliedin

100mMKPO4

1.0mMDTT

0.1mMEDTA

50%glycerol

pH6.5@25°C

SuppliedWith

B0110(10XBlueBuffer)

10XBlueBuffer(B0110)

500mMNaCl

100mMTris-HCl

100mMMgCl2

10mMDTT

pH7.9@25°C

PartNumber P7080L

Concentration 3,000U/ml

UnitSize 2,000U

StorageTemperature -15to-25°C

Purity(SDS-PAGE) >99%

SpecificActivity 5,555U/mg

E.coliDNAContamination 30U<10copies

自营商城图标
厂家直采
全球直采模式 正品优价
正品保障图标
正品保障
厂家直发 有线跟踪
解放采购图标
正规清关
CIF100%正规报关,提供发票
及时交付图标
及时交付
限时必达 不达必赔
在线客服
客服电话

4000-520-616

0512-67156496