LOCUS       sequence-129749-        8812 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 8812)
  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 8812)
  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-FLAG-DDB1
FEATURES             Location/Qualifiers
     source          1..8812
                     /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             890..913
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(4365..4383)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4366..4383)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4403..4420)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4409..4633
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4679..5107
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4766..4785)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4976..4997
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5116..5136)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5121..5450
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5301..5436
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5363..5382
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5517..6311
                     /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(5571..5590)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6181..6200
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6485..6606
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6522..6541)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6576..6595
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6655..6671)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6655..6671)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6668..6690)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6679..6695
                     /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(6703..6709,6710..6727,6728..6733))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6748..6769
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6886..6903)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7057..7645)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7137..7156)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7816..8607,8608..8676))
                     /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     8439..8458
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8677..8781)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct 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 atgggtggag 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 ggagacatgg actacaaaga
      901 cgatgacgat aaaccaagct tggtaccggg tatgtcgtac aactacgtgg taacggccca
      961 gaagcccacc gccgtgaacg gctgcgtgac cggacacttt acttcggccg aagacttaaa
     1021 cctgttgatt gccaaaaaca cgagattaga gatctatgtg gtcaccgccg aggggcttcg
     1081 gcccgtcaaa gaggtgggca tgtatgggaa gattgcggtc atggagcttt tcaggcccaa
     1141 gggggagagc aaggacctgc tgtttatctt gacagcgaag tacaatgcct gcatcctgga
     1201 gtataaacag agtggcgaga gcattgacat cattacgcga gcccatggca atgtccagga
     1261 ccgcattggc cgcccctcag agaccggcat tattggcatc attgaccctg agtgccggat
     1321 gattggcctg cgtctctatg atggcctttt caaggttatt ccactagatc gcgataataa
     1381 agaactcaag gccttcaaca tccgcctgga ggagctgcat gtcattgatg tcaagttcct
     1441 atatggttgc caagcaccta ctatttgctt tgtctaccag gaccctcagg ggcggcacgt
     1501 aaaaacctat gaggtgtctc tccgagaaaa ggaattcaat aagggccctt ggaaacagga
     1561 aaatgtcgaa gctgaagctt ccatggtgat cgcagtccca gagccctttg ggggggccat
     1621 catcattgga caggagtcaa tcacctatca caatggtgac aaatacctgg ctattgcccc
     1681 tcctatcatc aagcaaagca cgattgtgtg ccacaatcga gtggacccta atggctcaag
     1741 atacctgctg ggagacatgg aaggccggct cttcatgctg cttttggaga aggaggaaca
     1801 gatggatggc accgtcactc tcaaggatct ccgtgtagaa ctccttggag agacctctat
     1861 tgctgagtgc ttgacatacc ttgataatgg tgttgtgttt gtcgggtctc gcctgggtga
     1921 ctcccagctt gtgaagctca acgttgacag taatgaacaa ggctcctatg tagtggccat
     1981 ggaaaccttt accaacttag gacccattgt cgatatgtgc gtggtggacc tggagaggca
     2041 ggggcagggg cagctggtca cttgctctgg ggctttcaag gaaggttctt tgcggatcat
     2101 ccggaatgga attggaatcc acgagcatgc cagcattgac ttaccaggca tcaaaggatt
     2161 atggccactg cggtctgacc ctaatcgtga gactgatgac actttggtgc tctcttttgt
     2221 gggccagaca agagttctca tgttaaatgg agaggaggta gaagaaaccg aactgatggg
     2281 tttcgtggat gatcagcaga ctttcttctg tggcaacgtg gctcatcagc agcttatcca
     2341 gatcacttca gcatcggtga ggttggtctc tcaagaaccc aaagctctgg tcagtgaatg
     2401 gaaggagcct caggccaaga acatcagtgt ggcctcctgc aatagcagcc aggtggtggt
     2461 ggctgtaggc agggccctct actatctgca gatccatcct caggagctcc ggcagatcag
     2521 ccacacagag atggaacatg aagtggcttg cttggacatc accccattag gagacagcaa
     2581 tggactgtcc cctctttgtg ccattggcct ctggacggac atctcggctc gtatcttgaa
     2641 gttgccctct tttgaactac tgcacaagga gatgctgggt ggagagatca ttcctcgctc
     2701 catcctgatg accacctttg agagtagcca ttacctcctt tgtgccttgg gagatggagc
     2761 gcttttctac tttgggctca acattgagac aggtctgttg agcgaccgta agaaggtgac
     2821 tttgggcacc cagcccaccg tattgaggac ttttcgttct ctttctacca ccaacgtctt
     2881 tgcttgttct gaccgcccca ctgtcatcta tagcagcaac cacaaattgg tcttctcaaa
     2941 tgtcaacctc aaggaagtga actacatgtg tcccctcaat tcagatggct atcctgacag
     3001 cctggcgctg gccaacaata gcaccctcac cattggcacc atcgatgaga tccagaagct
     3061 gcacattcgc acagttcccc tctatgagtc tccaaggaag atctgctacc aggaagtgtc
     3121 ccagtgtttc ggggtcctct ccagccgcat tgaagtccaa gacacgagtg ggggcacgac
     3181 agccttgagg cccagcgcta gcacccaggc tctgtccagc agtgtaagct ccagcaagct
     3241 gttctccagc agcactgctc ctcatgagac ctcctttgga gaagaggtgg aggtgcacaa
     3301 cctacttatc attgaccaac acacctttga agtgcttcat gcccaccagt ttctgcagaa
     3361 tgaatatgcc ctcagtctgg tttcctgcaa gctgggcaaa gaccccaaca cttacttcat
     3421 tgtgggcaca gcaatggtgt atcctgaaga ggcagagccc aagcagggtc gcattgtggt
     3481 ctttcagtat tcggatggaa aactacagac tgtggctgaa aaggaagtga aaggggccgt
     3541 gtactctatg gtggaattta acgggaagct gttagccagc atcaatagca cggtgcggct
     3601 ctatgagtgg acaacagaga aggagctgcg cactgagtgc aaccactaca acaacatcat
     3661 ggccctctac ctgaagacca agggcgactt catcctggtg ggcgacctta tgcgctcagt
     3721 gctgctgctt gcctacaagc ccatggaagg aaactttgaa gagattgctc gagactttaa
     3781 tcccaactgg atgagtgctg tggaaatctt ggatgatgac aattttctgg gggctgaaaa
     3841 tgcctttaac ttgtttgtgt gtcaaaagga tagcgctgcc accactgacg aggagcggca
     3901 gcacctccag gaggttggtc ttttccacct gggcgagttt gtcaatgtct tttgccacgg
     3961 ctctctggta atgcagaatc tgggtgagac ttccaccccc acacaaggct cggtgctctt
     4021 cggcacggtc aacggcatga tagggctggt gacctcactg tcagagagct ggtacaacct
     4081 cctgctggac atgcagaatc gactcaataa agtcatcaaa agtgtgggga agatcgagca
     4141 ctccttctgg agatcctttc acaccgagcg gaagacagaa ccagccacag gtttcatcga
     4201 cggtgacttg attgagagtt tcctggatat tagccgcccc aagatgcagg aggtggtggc
     4261 aaacctacag tatgacgatg gcagcggtat gaagcgagag gccactgcag acgacctcat
     4321 caaggttgtg gaggagctaa ctcggatcct ctagagggcc ctattctata gtgtcaccta
     4381 aatgctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     4441 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     4501 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     4561 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     4621 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     4681 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     4741 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     4801 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     4861 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     4921 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     4981 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     5041 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     5101 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     5161 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     5221 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     5281 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     5341 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     5401 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     5461 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     5521 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     5581 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     5641 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     5701 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     5761 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     5821 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     5881 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     5941 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     6001 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     6061 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     6121 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     6181 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     6241 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     6301 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     6361 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     6421 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     6481 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     6541 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     6601 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     6661 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     6721 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     6781 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     6841 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     6901 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     6961 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     7021 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     7081 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     7141 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     7201 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     7261 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     7321 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     7381 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     7441 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     7501 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     7561 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     7621 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     7681 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     7741 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     7801 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     7861 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     7921 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     7981 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     8041 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8101 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     8161 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8221 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8281 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8341 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     8401 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8461 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8521 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8581 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     8641 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     8701 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     8761 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
