LOCUS       sequence-228469-        8912 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 8912)
  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 8912)
  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-HA2-DDB1
FEATURES             Location/Qualifiers
     source          1..8912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             899..925
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             929..955
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             935..961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     promoter        complement(4467..4485)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4468..4485)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4505..4522)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4511..4735
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4781..5209
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4868..4887)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5078..5099
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5218..5238)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5223..5552
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5403..5538
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5465..5484
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5619..6413
                     /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(5673..5692)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6283..6302
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6587..6708
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6624..6643)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6678..6697
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6757..6773)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6757..6773)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6770..6792)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6781..6797
                     /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(6805..6811,6812..6829,6830..6835))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6850..6871
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6988..7005)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7159..7747)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7239..7258)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7918..8709,8710..8778))
                     /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     8541..8560
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8779..8883)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctatgta
      901 cccatacgat gttcctgact atgcgggcta tccctatgac gtcccggact atgcaagctt
      961 ggtaccgtta acatggatcc gcatgtcgta caactacgtg gtaacggccc agaagcccac
     1021 cgccgtgaac ggctgcgtga ccggacactt tacttcggcc gaagacttaa acctgttgat
     1081 tgccaaaaac acgagattag agatctatgt ggtcaccgcc gaggggcttc ggcctgtcaa
     1141 agaggtgggc atgtatggga agattgcggt catggagctt ttcaggccca agggggagag
     1201 caaggacctg ctgtttatct tgacagcgaa gtacaatgcc tgcatcctgg agtataaaca
     1261 gagtggcgag agcattgaca tcattacgcg agcccatggc aatgtccagg accgcattgg
     1321 ccgcccctca gagaccggca ttattggcat cattgaccct gagtgccgga tgattggcct
     1381 gcgtctctat gatggccttt tcaaggttat tccactagat cgcgataata aagaactcaa
     1441 ggccttcaac atccgcctgg aggagctgca tgtcattgat gtcaagttcc tatatggttg
     1501 ccaagcacct actatttgct ttgtctacca ggaccctcag gggcggcacg taaaaaccta
     1561 tgaggtgtct ctccgagaaa aggaattcaa taagggccct tggaaacagg aaaatgtcga
     1621 agctgaagct tccatggtga tcgcagtccc agagcccttt gggggggcca tcatcattgg
     1681 acaggagtca atcacctatc acaatggtga caaatacctg gctattgccc ctcctatcac
     1741 caagcaaagc acgcttgtgt gccacaatcg agtggaccct aatggctcaa gatacctgct
     1801 gggagacatg gaaggccggc tcttcatgct gcttttggag aaggaggaac agatggatgg
     1861 caccgtcact ctcaaggatc tccgtgtaga actccttgga gagacctcta ttgctgagtg
     1921 cttgacatac cttgataatg gtgttgtgtt tgtcgggtct cgcctgggtg actcccagct
     1981 tgtgaagctc aacgttgaca gtaatgaaca aggctcctat gtagtggcca tggaaacctt
     2041 taccaactta ggacccattg tcgatatgtg cgtggtggac ctggagaggc aggggcaggg
     2101 gcagctggtc acttgctctg gggctttcaa ggaaggttct ttgcggatca tccggaatgg
     2161 aattggaatc cacgagcatg ccagcattga cttaccaggc atcaaaggat tatggccact
     2221 gcggtctgac cctaatcgtg agactgatga cactttggtg ctctcttttg tgggccagac
     2281 aagagttctc atgttaaatg gagaggaggt agaagaaacc gaactgatgg gtttcgtgga
     2341 tgatcagcag actttcttct gtggcaacgt ggctcatcag cagcttatcc agatcacttc
     2401 agcatcggtg aggttggtct ctcaagaacc caaagctctg gtcagtgaat ggaaggagcc
     2461 tcaggccaag aacatcagtg tggcctcctg caatagcagc caggtggtgg tggctgtagg
     2521 cagggccctc tactatctgc agatccatcc tcaggagctc cggcagatca gccacacaga
     2581 gatggaacat gaagtggctt gcttggacat caccccatta ggagacagca atggactgtc
     2641 ccctctttgt gccattggcc tctggacgga catctcggct cgtatcttga agttgccctc
     2701 ttttgaacta ctgcacaagg agatgctggg tggagagatc attcctcgct ccatcctgat
     2761 gaccaccttt gagagtagcc attacctcct ttgtgccttg ggagatggag cgcttttcta
     2821 ctttgggctc aacattgaga caggtctgtt gagcgaccgt aagaaggtga ctttgggcac
     2881 ccagcccacc gtattgagga cttttcgttc tctttctacc accaacgtct ttgcttgttc
     2941 tgaccgcccc actgtcatct atagcagcaa ccacaaattg gtcttctcaa atgtcaacct
     3001 caaggaagtg aactacgtgt gtcccctcaa ttcagatggc tatcctgaca gcctggcgct
     3061 ggccaacaat agcaccctca ccattggcac catcgatgag atccagaagc tgcacattcg
     3121 cacagttccc ctctatgagt ctccaaggaa gatctgctac caggaagtgt cccagtgttt
     3181 cggggtcctc tccagccgca ttgaagtcca agacacgagt gggggcacga cagccttgag
     3241 gcccagcgct agcacccagg ctctgtccag cagtgtaagc tccagcaagc tgttctccag
     3301 cagcactgct cctcatgaga cctcctttgg agaagaggtg gaggtgcaca acctacttat
     3361 cattgaccaa cacacctttg aagtgcttca tgcccaccag tttctgcaga atgaatatgc
     3421 cctcagtctg gtttcctgca agctgggcaa agaccccaac acttacttca ttgtgggcac
     3481 agcaatggtg tatcctgaag aggcagagcc caagcagggt cgcattgtgg tctttcagta
     3541 ttcggatgga aaactacaga ctgtggctga aaaggaagtg aaaggggccg tgtactctat
     3601 ggtggaattt aacgggaagc tgttagccag catcaatagc acggtgcggc tctatgagtg
     3661 gacaacagag aaggagctgc gcactgagtg caaccactac aacaacatca tggccctcta
     3721 cctgaagacc aagggcgact tcatcctggt gggcgacctt atgcgctcag tgctgctgct
     3781 tgcctacaag cccatggaag gaaactttga agagattgct cgagacttta atcccaactg
     3841 gatgagtgct gtggaaatct tggatgatga caattttctg ggggctgaaa atgcctttaa
     3901 cttgtttgtg tgtcaaaagg atagcgctgc caccactgac gaggagcggc agcacctcca
     3961 ggaggttggt cttttccacc tgggcgagtt tgtcaatgtc ttttgccacg gctctctggt
     4021 aatgcagaat ctgggtgaga cttccacccc cacacaaggc tcggtgctct tcggcacggt
     4081 caacggcatg atagggctgg tgacctcact gtcagagagc tggtacaacc tcctgctgga
     4141 catgcagaat cgactcaata aagtcatcaa aagtgtgggg aagatcgagc actccttctg
     4201 gagatccttt cacaccgagc ggaagacaga accagccaca ggtttcatcg acggtgactt
     4261 gattgagagt ttcctggata ttagccgccc caagatgcag gaggtggtgg caaacctaca
     4321 gtatgacgat ggcagcggta tgaagcgaga ggccactgca gacgacctca tcaaggttgt
     4381 ggaggagcta actcggattc atacgtaggc tagcgtcgac aagggcgaat tccagcacac
     4441 tggcggccgt tactagaggg ccctattcta tagtgtcacc taaatgctag agctcgctga
     4501 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     4561 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     4621 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     4681 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     4741 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     4801 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     4861 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     4921 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     4981 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     5041 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     5101 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     5161 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     5221 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     5281 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     5341 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     5401 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     5461 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     5521 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     5581 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     5641 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     5701 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     5761 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     5821 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     5881 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     5941 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     6001 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     6061 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     6121 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     6181 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     6241 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     6301 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     6361 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     6421 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     6481 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     6541 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     6601 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     6661 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     6721 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     6781 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     6841 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     6901 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6961 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     7021 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     7081 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     7141 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     7201 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     7261 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     7321 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     7381 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     7441 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     7501 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     7561 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     7621 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     7681 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     7741 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     7801 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     7861 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     7921 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     7981 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8041 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     8101 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8161 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8221 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8281 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8341 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8401 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8461 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8521 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8581 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8641 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8701 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8761 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     8821 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     8881 gcgcacattt ccccgaaaag tgccacctga cg
//
