LOCUS       sequence-43811-e        6549 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 6549)
  AUTHORS   Angers S, Thorpe CJ, Biechele TL, Goldenberg SJ, Zheng N, MacCoss 
            MJ, Moon RT
  TITLE     The KLHL12-Cullin-3 ubiquitin ligase negatively regulates the 
            Wnt-beta-catenin pathway by targeting Dishevelled for degradation.
  JOURNAL   Nat Cell Biol. 2006 Apr . 8(4):348-57.
  PUBMED    16547521
REFERENCE   2  (bases 1 to 6549)
  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: "Nat Cell 
            Biol. 2006 Apr . 8(4):348-57."
COMMENT     Sequence Label: XE256 pCDNA3.1(zeo)-hDsh3-deltaCTER
FEATURES             Location/Qualifiers
     source          1..6549
                     /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"
     regulatory      935..944
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             944..967
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    1592..1603
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     primer_bind     complement(2556..2573)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2562..2786
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2832..3260
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2919..2938)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3129..3150
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3269..3289)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3274..3604
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3455..3590
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3517..3536
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3652..3699
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3718..4092
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4222..4343
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4259..4278)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4313..4332
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4392..4408)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4392..4408)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4405..4427)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4416..4432
                     /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(4440..4446,4447..4464,4465..4470))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4485..4506
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4623..4640)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4794..5382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4874..4893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5553..6344,6345..6413))
                     /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     6176..6195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6414..6518)
                     /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 gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccgccacc atggactaca aggacgacga
      961 cgacaagggc ggcgcgcctg tacagattcg agagggtgga agtggaccag gtggtatggg
     1021 cgagaccaag atcatctacc acttggatgg gcaggagacg ccgtaccttg tgaagctgcc
     1081 cctgcccgcc gagcgcgtca ccttggcgga ctttaagggc gttttgcagc gacccagcta
     1141 taagttcttc ttcaagtcta tggacgacga tttcggagtg gtgaaggagg agatctcgga
     1201 tgacaatgcc aagctaccat gcttcaatgg ccgggtggtg tcctggctgg tgtcagctga
     1261 gggctcacac ccagacccag cccccttctg tgctgataac ccatcggagc tgccaccacc
     1321 tatggagcgc acgggaggca tcggggactc ccgaccccca tccttccacc ctcatgctgg
     1381 tgggggcagc caggagaacc tggacaatga cacagagacg gactctttgg tgtctgccca
     1441 gcgagagcgg ccacgccgga gggatggccc agagcatgca acccggctaa atggaactgc
     1501 gaagggggaa cggcggcgag aaccaggggg ttatgatagc tcatccaccc ttatgagcag
     1561 tgagctggag accaccagct tctttgactc agatgaggat gactccacca gcaggttcag
     1621 cagctccaca gaacagagca gtgcctcacg cctgatgaga agacacaagc ggcggcggcg
     1681 gaagcagaag gtttctcgga ttgagcggtc ctcgtccttc agcagcatca cggactccac
     1741 catgtcactc aacatcatca cggtcactct caacatggaa aaatataact tcttgggcat
     1801 ctccattgtg ggccaaagca acgagcgtgg tgacggcggc atctacattg gctctatcat
     1861 gaagggtggg gccgtggctg ctgatggacg catcgagcca ggagatatgt tgttacaggt
     1921 aaacgagatc aactttgaga acatgagtaa tgacgatgca gtccgggtac tgcgggagat
     1981 tgtgcacaaa ccggggccca tcaccctgac tgtagccaag tgctgggacc caagtccacg
     2041 tggttgcttc acattgccca ggagcgagcc catccggccc attgaccctg cggcctgggt
     2101 ctcccacact gcagccatga ccggcacctt ccctgcatac ggcatgagcc cctccctgag
     2161 caccatcacc tccaccagct cctccatcac cagttccatc cctgacacag agcgcctaga
     2221 cgacttccac ttgtccatcc acagtgacat ggctgccatc gtaaaagcca tggcctcccc
     2281 tgaatcaggg ttggaggtcc gtgaccgcat gtggctcaag attaccatcc ctaatgcttt
     2341 catcggctca gatgtggtgg actggctgta ccacaatgtg gaaggcttca cggaccggag
     2401 ggaggcccgc aagtatgcca gcaacctgct gaaagctggc ttcatccgcc ataccgtcaa
     2461 caagatcacc ttctccgagc agtgctacta catcttcggt taggaattcg cccttttcgg
     2521 atccactaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
     2581 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     2641 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     2701 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     2761 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag
     2821 ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2881 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2941 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcggggca tccctttagg
     3001 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     3061 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     3121 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     3181 ttttgattta taagggattt tggggatttc ggcctattgg ttaaaaaatg agctgattta
     3241 acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3301 ccaggctccc caggcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     3361 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     3421 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     3481 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     3541 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     3601 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
     3661 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
     3721 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
     3781 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
     3841 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
     3901 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
     3961 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
     4021 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
     4081 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     4141 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     4201 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     4261 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4321 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     4381 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     4441 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4501 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4561 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     4621 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     4681 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     4741 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4801 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4861 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4921 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4981 ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5041 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5101 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5161 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5221 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5281 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5341 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5401 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5461 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5521 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     5581 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     5641 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     5701 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     5761 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5821 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5881 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5941 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6001 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6061 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6121 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6181 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6241 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6301 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6361 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6421 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6481 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6541 cctgacgtc
//
