- Convenient,pre-processedvectorseliminatetheneedfordigestion,gelpurification,anddephosphorylation.
- TranscriptionterminatorsflankingcloningsitestABIlizeotherwisetoxicinserts.
- Choiceofcopynumberandantibioticresistance.
- OptimizedforusewithE.cloni®10Gelectrocompetentandchemicallycompetentcells(soldseparately)
Clonequicklyandfacilitatedownstreamapplications
Lucigen’suniquecloningkitswithpre-processedvectorsandhighlycompetentcellseliminatehoursoftediouspreparationandenableefficientcloning.Ligationcanbecompletedinaslittleas30minuteswithnoclean-upneeded;onlyabriefheatdenaturationisnecessarybeforetransformation.Downstreammanipulation(e.g,mutagenesis,subsequentinsertaddition)isfacilitatedbytheminimalvectorsize(1.7-2.0kb).
StabilizedInsertsandGap-FreeCloning
Allcloningkitsincludethetranscription-andtranslation-freepSMART®vectors(Figure1),whicheliminatemanyoftheproblemsassociatedwithcloningrecalcitrantDNAinconventionalplasmidvectors.Conventionalcloningvectorscontainpromoters(e.g.,lacZpromoter)thatconstitutivelytranscribetheinsertsequence.Thistranscriptionandcoupledtranslationcandestabilizeinsertsthatcontaincodingregions,strongpromoters,shortrepeats,orincompatIBLesecondarystructures.ThecloningsiteofpSMARTvectorsdoesnotincludealaczsequence.Inaddition,strongtranscriptionterminatorsflankthecloningsitetoblockspurioustranscriptionfromthevectorandinsert-driventranscriptionintothevector.LowcopynumberversionsofpSMARTfurtherstabilizesequencesthataredifficulttomaintainintypicalvectors.
NoBackgroundProblems
ThepSMARTvectorsaresuppliedpre-digested,withblunt,dephosphorylatedends,andarequalifiedtoproduce99.5%recombinantclonesintypicalexperiments.Theultra-lowbackgroundofemptyvector(lessthan0.5%)eliminatestheneedtoscreenforrecombinants,removestheuncertaintyoffalsenegatives(lightbluepUCcolonies)andfalsepositives(whitecoloniesthatlackinserts),andenableslibraryconstructionfromnanogramamountsofDNA.
Figure1.Highcopy(HC)andlowcopy(LC)versionsofthepSMARTtranscription-freecloningvectors.
ConvenientSuccess
EfficientlyobtainyourcloneorcreateyourgenomicorCDNAlibrary—theCloneSmartCloningKitseliminatetediouspreparationofvectorsandcompetentcells,aswellastime-consumingQCtestingexperiments.Kitsincludeoptimizedreagents,ligation-readyvector(nopost-ligationcleanupsteprequired),detailedinstructions,andtrouble-shootingguidestosimplifycloningandsequencing.HighlyefficientE.cloni®10Gelectrocompetentcells(upto>4×1010cfu/µg)orchemicallycompetentcells(>1×109cfu/µg)areavailableforsaleseparately.
ORDERINFORMATION
CloneSmartBluntCloningKitscontain:VectorPremix,CloneSmartDNALigase,PositiveControlInsertDNA,SequencingPrimers,andacompleteprotocol.KitscanbeusedwithanyE.colichemicallycompetentorelectrocompetentcellsbuthavebeenoptimizedforusewithofE.cloni®ElectrocompetentCellsandChemicallyCompetentCells.ebiomall.com
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般基克隆物体直接取基包括面几步骤
提取植物或者物基组DNA（试剂盒）————做PCR需要基扩增（顺利2搞定）——扩增基做酶切——基连接想插入载体面（顺利1-2）——鉴定克隆基
步骤都商品化试剂或者试剂盒 要思路清晰做起快 1-2周能完
没证实情况任何没依据推断都假想空想
现在小弟碰见一个难题，我想克隆一个基因，但是我在NCBI一检索，发现别人只发表了这个基因的DNA全长，但是，CDNA没有发表，也没有发表文章，想请问下，在不知道CDNA的情况下，请问我应该如何做这个基因的克隆？？？（全基因组也不知道,其他生物没有这种基因）
详见：http://zhidao.baidu.com/question/1831844001436785260.html?fr=iks&word=%B2%A1%B6%BE%B5%C4%B8%B4%D6%C6%D6%DC%C6%DA%B5%C4%B9%FD%B3%CC%3F&ie=gbk
