pET28a-c()质粒图谱.pdf

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1、pET-28a-c(+)VectorsTB07412/98Cat.No.ThepET-28a-c(+)vectorscarryanN-terminalHis•Tag®/thrombin/T7•Tag®configurationpluspET-28aDNA69864-3anoptionalC-terminalHis•Tagsequence.Uniquesitesareshownonthecirclemap.NotethatthepET-28bDNA69865-3sequenceisnumberedbythepBR322convention,sotheT7express

2、ionregionisreversedonthepET-28cDNA69866-3circularmap.Thecloning/expressionregionofthecodingstrandtranscribedbyT7RNApoly-Call137-649-32037tobuythevectorinChina.meraseisshownbelow.Thef1originisorientedsothatinfectionwithhelperphagewillproducevirionscontainingsingle-strandedDNAthatcorres

3、pondstothecodingstrand.Therefore,single-strandedsequencingshouldbeperformedusingtheT7terminatorprimer(Cat.No.69337-3).XhoI(158)NotI(166)pET-28a(+)sequencelandmarksEagI(166)HindIII(173)T7promoter370-386SalI(179)T7transcriptionstart369SacI(190)EcoRI(192)His•Tagcodingsequence270-287BamHI

4、(198)T7•Tagcodingsequence207-239Bpu1102I(80)NheI(231)NdeI(238)MultiplecloningsitesDraIII(5127)NcoI(296)(BamHI-XhoI)158-203XbaI(335)His•Tagcodingsequence140-157BglII(401)03-5358)SgrAI(442)T7terminator26-72in(49SphI(598)riglacIcodingsequence773-1852of1pBR322origin3286)Kancodingsequence3

5、995-4807PvuI(4426)70f1origin4903-5358SgfI(4426)84-5SmaI(4300)99MluI(1123)3ThemapsforpET-28b(+)andpET-28c(+)(lacBclI(1137)narethesameaspET-28a(+)(shown)withIaClaI(4117)(thefollowingexceptions:pET-28b(+)isaK77BstEII(1304)NruI(4083)35368bpplasmid;subtract1bpfromeachsite1-pET-28a(+)8ApaI(

6、1334)beyondBamHIat198.pET-28c(+)isa(5369bp)525367bpplasmid;subtract2bpfromeachsite)BssHII(1534)beyondBamHIat198.Eco57I(3772)EcoRV(1573)HpaI(1629)AlwNI(3640)ori(3286PshAIBssSI(3397))(1968)BspLU11I(3224)BglI(2187)SapI(3108)FspI(2205)Bst1107I(2995)Psp5II(2230)Tth111I(2969)T7promoterprime

7、r#69348-3pETupstreamprimer#69214-3BglIIT7promoterlacoperatorXbaIrbsAGATCTCGATCCCGCGAAATTAATACGACTCACTATAGGGGAATTGTGAGCGGATAACAATTCCCCTCTAGAAATAATTTTGTTTAACTTTAAGAAGGAGANcoIHis•TagNdeINheIT7•TagTATACCATGGGCAGCAGCCATCATCATCATCATCACAGCAGCGGCCTGGTGCCGCGCGGCAGCCATATGGCTAGCATGACTGGTGGACAGCA

8、AMetG