LOCUS       sequence-9415-ex        7113 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7113)
  AUTHORS   Hu J, Zacharek S, He YJ, Lee H, Shumway S, Duronio RJ, Xiong Y
  TITLE     WD40 protein FBW5 promotes ubiquitination of tumor suppressor TSC2 
            by DDB1-CUL4-ROC1 ligase.
  JOURNAL   Genes Dev. 2008 Apr 1. 22(7):866-71.
  PUBMED    18381890
REFERENCE   2  (bases 1 to 7113)
  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"; journalName: "Genes Dev. 
            2008 Apr 1. 22(7):866-71."
COMMENT     Sequence Label: pcDNA3-FBL6-FLAG3C
FEATURES             Location/Qualifiers
     source          1..7113
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2588..2653
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        complement(2665..2683)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2666..2683)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2703..2720)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2709..2933
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2979..3407
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3066..3085)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3276..3297
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3416..3436)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3421..3751
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3602..3737
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3664..3683
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3818..4612
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3872..3891)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4482..4501
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4786..4907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4823..4842)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4877..4896
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4956..4972)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4956..4972)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4969..4991)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4980..4996
                     /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)."
     promoter        complement(join(5004..5010,5011..5028,5029..5034))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5049..5070
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5187..5204)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5358..5946)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5438..5457)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6117..6908,6909..6977))
                     /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     6740..6759
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6978..7082)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcatggctg ccccagcctc
      961 ccggcaggtc cgacgcagag cccgggcagc gccgcggccc cgctcggccg aggactggtg
     1021 gtgggaccgg ctggcgccga ggggctcggg gtaccacctg ctgcagtccg acagcatgct
     1081 gctggtgctg tccgaacccg gccccgcccg gccccgcgca cagcggcgcg cttcccgccg
     1141 cactccccgg cagccgcccc ggggccccag cgccgcggcc aagcccaagg ccgggctcag
     1201 gtccgaggcg gcggccgcgc ccgcacccgc accggcaccc acgcccacgc ccgaggaagg
     1261 gcccgacgcg ggctggggag accgcattcc cttggaaatc ctggtgcaga ttttcgggtt
     1321 gttggtggcg gcggacggcc ccatgccctt cctgggcagg gctgcgcgcg tgtgccgccg
     1381 ctggcaggag gccgcttccc aacccgcgct ctggcacacc gtgaccctgt cgtccccgct
     1441 ggtcggccgg cctgccaagg gcggggtcaa ggcggagaag aagctccttg cttccctgga
     1501 gtggcttatg cccaatcggt tttcacagct ccagaggctg accctcatcc actggaagtc
     1561 tcaggtacac cccgtgttga agctggtagg tgagtgctgt cctcggctca ctttcctcaa
     1621 gctctccggc tgccacggtg tgactgctga cgctctggtc atgctagcca aagcctgctg
     1681 ccagctccat agcctggacc tacagcactc catggtggag tccacagctg tggtgagctt
     1741 cttggaggag gcagggtccc gaatgcgcaa gttgtggctg acctacagct cccagacgac
     1801 agccatcctg ggcgcactgc tgggcagctg ctgcccccag ctccaggtcc tggaggtgag
     1861 caccggcatc aaccgtaata gcattcccct tcagctgcct gtcgaggctc tgcagaaagg
     1921 ctgccctcag ctccaggtgc tgcggctgtt gaacctgatg tggctgccca agcctccggg
     1981 acgaggggtg gctcccggac caggcttccc tagcctagag gagctctgcc tggcgagctc
     2041 aacctgcaac tttgtgagca acgaggtcct gggccgccta ctccacggct ctcccaacct
     2101 gcgcttactg gatcttcgtg gctgtgcgcg catcacgccg gctggccttc aggatctgcc
     2161 atgtcgggag ctggagcagc ttcatctggg cctgtatggc acgtcagacc ggctgactct
     2221 agccaaggag ggcagcccct ttttgaccca gaagtggtgc catacactgc gagaactgga
     2281 cttgagtggc caggggttca gtgagaagga cctggagcag gccctggctg ccttcttaag
     2341 cacccctggg ggctcacacc cagccctgtg ctctcttaac ctcaggggca cccgggtcac
     2401 accaagcact gtcagctctg tgatcagcag ctgcccgggc ctgctctacc tcaacctgga
     2461 gtcctgccgc tgccttcccc ggggtctgaa gcgggcctac cggggcctgg aggaagtcca
     2521 gtggtgtctg gagcagctgc tcaccagccc ctcacccagc tcgagcatgc atctagagtt
     2581 aaccatggac tacaaagacc atgacggtga ttataaagat catgatatcg attacaagga
     2641 tgacgatgac aagtagggcc ctattctata gtgtcaccta aatgctagag ctcgctgatc
     2701 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     2761 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     2821 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     2881 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     2941 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3001 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3061 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3121 agctctaaat cggggcatcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3181 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3241 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3301 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgg ggatttcggc
     3361 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3421 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag gcaggcagaa gtatgcaaag
     3481 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     3541 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     3601 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     3661 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg
     3721 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
     3781 cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     3841 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     3901 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     3961 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     4021 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     4081 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     4141 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     4201 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     4261 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     4321 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     4381 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     4441 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     4501 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     4561 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     4621 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     4681 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     4741 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca
     4801 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     4861 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata
     4921 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     4981 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     5041 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     5101 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     5161 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     5221 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5281 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5341 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     5401 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5461 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     5521 tttctccctt cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg
     5581 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     5641 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     5701 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     5761 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     5821 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     5881 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     5941 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     6001 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     6061 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     6121 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     6181 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     6241 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     6301 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     6361 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     6421 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     6481 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     6541 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     6601 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     6661 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     6721 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     6781 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     6841 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     6901 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     6961 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     7021 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     7081 cgcacatttc cccgaaaagt gccacctgac gtc
//
