LOCUS       sequence-8522-ex        7212 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 7212)
  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 7212)
  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: XE251 pCDNA3.1(zeo)-hDsh3
FEATURES             Location/Qualifiers
     source          1..7212
                     /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(3219..3236)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3225..3449
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3495..3923
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3582..3601)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3792..3813
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3932..3952)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3937..4267
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4118..4253
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4180..4199
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4315..4362
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4381..4755
                     /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    4885..5006
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4922..4941)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4976..4995
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5055..5071)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5055..5071)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5068..5090)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5079..5095
                     /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(5103..5109,5110..5127,5128..5133))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5148..5169
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5286..5303)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5457..6045)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5537..5556)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6216..7007,7008..7076))
                     /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     6839..6858
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7077..7181)
                     /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 gacctctgcg gcaacatggc
     2521 caacctgtct ctccacgatc acgatggctc cagtggcgcc tctgaccagg acacactggc
     2581 ccctttgccg cacccggggg ccgccccttg gcccatggct ttcccgtacc agtacccgcc
     2641 acgcccgcac ccatacaacc cgcacccggg cttgggggag ctgggctaca gctacggcgg
     2701 gggcagcgcc agcagtcagc acagcgaagg cagtcggagc agtggctcca accgtagcgg
     2761 cagcgatcgg aggaaggaga aggacccgaa ggccggggac tccaagtccg ggggcagcgg
     2821 cagcgaatcg gaccacacca cacgcagcag cctgcggggg ccgcgggagc gggcgcccag
     2881 cgagcgctca gggccggcgg ccagcgagca cagccaccgc agccaccatt ccctggccag
     2941 cagccttcgc agccaccaca cacacccgag ctacggtcct cccggagtgc cccctctcta
     3001 cggccccccc atgctgatga tgcccccgcc gcccgcggcc atggggcccc caggagcccc
     3061 tccgggccgc gacctggcct cagtgccccc ggaactgacc gccagcagac agtccttccg
     3121 catggccatg ggaaacccca gtgagttctt tgtggatgtg atgtgagaat tcgccctttt
     3181 cggatccact agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     3241 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     3301 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     3361 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     3421 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     3481 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     3541 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     3601 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg gcatcccttt
     3661 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     3721 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     3781 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     3841 ttcttttgat ttataaggga ttttggggat ttcggcctat tggttaaaaa atgagctgat
     3901 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     3961 tccccaggct ccccaggcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4021 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4081 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     4141 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     4201 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     4261 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
     4321 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     4381 atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc
     4441 gagttctgga ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt
     4501 gtggtccggg acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac
     4561 aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag
     4621 gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag
     4681 ccgtgggggc gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc
     4741 gaggagcagg actgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4801 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4861 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     4921 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     4981 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     5041 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     5101 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     5161 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5221 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     5281 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5341 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5401 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5461 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5521 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5581 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     5641 gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     5701 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     5761 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     5821 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     5881 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     5941 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     6001 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6061 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6121 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     6181 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6241 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6301 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6361 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6421 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6481 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6541 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6601 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     6661 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     6721 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     6781 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     6841 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6901 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6961 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     7021 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7081 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7141 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7201 ccacctgacg tc
//
