LOCUS       sequence-173591-        7364 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Gateway-compatible expression vector backbone with an 
            N-terminal 3xFLAG-V5 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7364)
  AUTHORS   Taipale M, Krykbaeva I, Koeva M, Kayatekin C, Westover KD, Karras 
            GI, Lindquist S
  TITLE     Quantitative analysis of HSP90-client interactions reveals 
            principles of substrate recognition.
  JOURNAL   Cell. 2012 Aug 31;150(5):987-1001. doi: 10.1016/j.cell.2012.06.047.
  PUBMED    22939624
REFERENCE   2  (bases 1 to 7364)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2012.06"; journalName: "Cell"; date: "2012-08-31- 
            31"; volume: "150"; issue: "5"; pages: "987-1001"
COMMENT     Sequence Label: pcDNA3.1-3xFLAG-V5-ccdB
FEATURES             Location/Qualifiers
     source          1..7364
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     181..201
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(210..638)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(320..341)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     532..551
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    684..908
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     897..914
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     protein_bind    965..1089
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1130..1435)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(1232..1251)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1777..2436)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2351..2370
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(2490..2496,2497..2514,2515..2520))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(2545..2669)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2667..2708)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(2727..2792)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2861..2885)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2882..3085)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2911..2931)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3086..3465
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     3637..3656
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3731..3835
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3836..3904,3905..4696)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4054..4073)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4867..5455
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5356..5375
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5609..5626
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5743..5764
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5779..5784,5785..5802,5803..5809)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5817..5833
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5822..5844
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5841..5857
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5841..5857
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    5906..6027
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5917..5936)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5971..5990
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6157..7182)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(6157..7182)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     promoter        complement(join(7231..7364,1..196))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(7245..7364,1..16)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7299..7318)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gagttagggg cgggatgggc ggagttaggg gcgggactat ggttgctgac taattgagat
       61 gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac ctggttgctg
      121 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
      181 accctaactg acacacattc cacagaatta attcgcgtta aatttttgtt aaatcagctc
      241 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      301 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      361 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      421 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      481 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      541 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      601 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtgg ggataccccc tagagcccca
      661 gctggttctt tccgcctcag aagccataga gcccaccgca tccccagcat gcctgctatt
      721 gtcttcccaa tcctccccct tgctgtcctg ccccacccca ccccccagaa tagaatgaca
      781 cctactcaga caatgcgatg caatttcctc attttattag gaaaggacag tgggagtggc
      841 accttccagg gtcaaggaag gcacggggga ggggcaaaca acagatggct ggcaactaga
      901 aggcacagtc gaggctgatc agcgggttta aacgggccct ctagactcga gttactatag
      961 agcaaccact ttgtacaaga aagctgaacg agaaacgtaa aatgatataa atatcaatat
     1021 attaaattag attttgcata aaaaacagac tacataatac tgtaaaacac aacatatcca
     1081 gtcactatgg tcgacctgca gactggctgt gtataaggga gcctgacatt tatattcccc
     1141 agaacatcag gttaatggcg tttttgatgt cattttcgcg gtggctgaga tcagccactt
     1201 cttccccgat aacggagacc ggcacactgg ccatatcggt ggtcatcatg cgccagcttt
     1261 catccccgat atgcaccacc gggtaaagtt cacgggagac tttatctgac agcagacgtg
     1321 cactggccag ggggatcacc atccgtcgcc cgggcgtgtc aataatatca ctctgtacat
     1381 ccacaaacag acgataacgg ctctctcttt tataggtgta aaccttaaac tgcatttcac
     1441 cagcccctgt tctcgtcagc aaaagagccg ttcatttcaa taaaccgggc gacctcagcc
     1501 atcccttcct gattttccgc tttccagcgt tcggcacgca gacgacgggc ttcattctgc
     1561 atggttgtgc ttaccagacc ggagatattg acatcatata tgccttgagc aactgatagc
     1621 tgtcgctgtc aactgtcact gtaatacgct gcttcatagc atacctcttt ttgacatact
     1681 tcgggtatac atatcagtat atattcttat accgcaaaaa tcagcgcgca aatacgcata
     1741 ctgttatctg gcttttagta agccggatcc agatctttac gccccgccct gccactcatc
     1801 gcagtactgt tgtaattcat taagcattct gccgacatgg aagccatcac aaacggcatg
     1861 atgaacctga atcgccagcg gcatcagcac cttgtcgcct tgcgtataat atttgcccat
     1921 ggtgaaaacg ggggcgaaga agttgtccat attggccacg tttaaatcaa aactggtgaa
     1981 actcacccag ggattggctg agacgaaaaa catattctca ataaaccctt tagggaaata
     2041 ggccaggttt tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa actgccggaa
     2101 atcgtcgtgg tattcactcc agagcgatga aaacgtttca gtttgctcat ggaaaacggt
     2161 gtaacaaggg tgaacactat cccatatcac cagctcaccg tctttcattg ccatacggaa
     2221 ttccggatga gcattcatca ggcgggcaag aatgtgaata aaggccggat aaaacttgtg
     2281 cttatttttc tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg tctggttata
     2341 ggtacattga gcaactgact gaaatgcctc aaaatgttct ttacgatgcc attgggatat
     2401 atcaacggtg gtatatccag tgattttttt ctccatttta gcttccttag ctcctgaaaa
     2461 tctcgacgga tcctaactca aaatccacac attatacgag ccggaagcat aaagtgtaaa
     2521 gcctggggtg cctaatgcgg ccgccatagt gactggatat gttgtgtttt acagtattat
     2581 gtagtctgtt ttttatgcaa aatctaattt aatatattga tatttatatc attttacgtt
     2641 tctcgttcag cttttttgta caaacttgta gaatcgagac cgaggagagg gttagggata
     2701 ggcttacccg aggtcgagcc cgagcccttg tcatcgtcat ccttgtagtc gatgtcatga
     2761 tctttataat caccgtcatg gtctttgtag tccatggctg gatcctctag agtcggtgtc
     2821 ttctatggag gtcaaaacag cgtggatggc gtctccaggc gatctgacgg ttcactaaac
     2881 gagctctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg
     2941 cggagttgtt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat
     3001 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat
     3061 tgatgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact
     3121 gccaagtagg aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac
     3181 cgtcattgac gtcaataggg ggcgtacttg gcatatgata cacttgatgt actgccaagt
     3241 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta
     3301 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag
     3361 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg
     3421 accccgtaat tgattactat taataactag tcaataatca atgtcaacgc gtatatctgg
     3481 cccgtacatc gcgaagcagc gcaaaacgcc taaccctaag cagattcttc atgcaattgt
     3541 cggtcaagcc ttgccttgtt gtagcttaaa ttttgctcgc gcactactca gcgacctcca
     3601 acacacaagc agggagcaga tactggctta actatgcggc atcagagcag attgtactga
     3661 gagtgcacca taggggatcg ggagatctcc cgatccgtcg acgtcaggtg gcacttttcg
     3721 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3781 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     3841 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     3901 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     3961 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     4021 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4081 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4141 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4201 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4261 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4321 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4381 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4441 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4501 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4561 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4621 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4681 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     4741 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     4801 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     4861 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     4921 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     4981 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5041 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5101 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5161 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagtg
     5221 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5281 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5341 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5401 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5461 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5521 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5581 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     5641 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     5701 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     5761 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     5821 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agctctagct
     5881 agaggtcgac ggtatacaga catgataaga tacattgatg agtttggaca aaccacaact
     5941 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     6001 accattataa gctgcaataa acaagttggg gtgggcgaag aactccagca tgagatcccc
     6061 gcgctggagg atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata
     6121 gaaggcggcg gtggaatcga aatctcgtag cacgtgctat tcctttgccc tcggacgagt
     6181 gctggggcgt cggtttccac tatcggcgag tacttctaca cagccatcgg tccagacggc
     6241 cgcgcttctg cgggcgattt gtgtacgccc gacagtcccg gctccggatc ggacgattgc
     6301 gtcgcatcga ccctgcgccc aagctgcatc atcgaaattg ccgtcaacca agctctgata
     6361 gagttggtca agaccaatgc ggagcatata cgcccggagc cgcggcgatc ctgcaagctc
     6421 cggatgcctc cgctcgaagt agcgcgtctg ctgctccata caagccaacc acggcctcca
     6481 gaagaagatg ttggcgacct cgtattggga atccccgaac atcgcctcgc tccagtcaat
     6541 gaccgctgtt atgcggccat tgtccgtcag gacattgttg gagccgaaat ccgcgtgcac
     6601 gaggtgccgg acttcggggc agtcctcggc ccaaagcatc agctcatcga gagcctgcgc
     6661 gacggacgca ctgacggtgt cgtccatcac agtttgccag tgatacacat ggggatcagc
     6721 aatcgcgcat atgaaatcac gccatgtagt gtattgaccg attccttgcg gtccgaatgg
     6781 gccgaacccg ctcgtctggc taagatcggc cgcagcgatc gcatccatgg cctccgcgac
     6841 cggctgcaga acagcgggca gttcggtttc aggcaggtct tgcaacgtga caccctgtgc
     6901 acggcgggag atgcaatagg tcaggctctc gctgaattcc ccaatgtcaa gcacttccgg
     6961 aatcgggagc gcggccgatg caaagtgccg ataaacataa cgatctttgt agaaaccatc
     7021 ggcgcagcta tttacccgca ggacatatcc acgccctcct acatcgaagc tgaaagcacg
     7081 agattcttcg ccctccgaga gctgcatcag gtcggagacg ctgtcgaact tttcgatcag
     7141 aaacttctcg acagacgtcg cggtgagttc aggctttttc atcacgtgct gatcagatcc
     7201 gaaaatggat atacaagctc ccgggagctt tttgcaaaag cctaggcctc caaaaaagcc
     7261 tcctcactac ttctggaata gctcagaggc agaggcggcc tcggcctctg cataaataaa
     7321 aaaaattagt cagccatggg gcggagaatg ggcggaactg ggcg
//
