LOCUS       sequence-8540-ex        6330 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 6330)
  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 6330)
  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: XE257 pCDNA3.1(zeo)-hDsh3-truncDEP
FEATURES             Location/Qualifiers
     source          1..6330
                     /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(2337..2354)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2343..2567
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2613..3041
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2700..2719)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2910..2931
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3050..3070)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3055..3385
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3236..3371
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3298..3317
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3433..3480
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3499..3873
                     /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    4003..4124
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4040..4059)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4094..4113
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4173..4189)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4173..4189)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4186..4208)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4197..4213
                     /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(4221..4227,4228..4245,4246..4251))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4266..4287
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4404..4421)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4575..5163)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4655..4674)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5334..6125,6126..6194))
                     /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     5957..5976
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6195..6299)
                     /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 ttaggaattc gcccttttcg gatccactag agggcccgtt taaacccgct gatcagcctc
     2341 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     2401 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     2461 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     2521 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     2581 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     2641 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2701 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2761 aaatcggggc atccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2821 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2881 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2941 caaccctatc tcggtctatt cttttgattt ataagggatt ttggggattt cggcctattg
     3001 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     3061 cagttagggt gtggaaagtc cccaggctcc ccaggcaggc agaagtatgc aaagcatgca
     3121 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3181 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3241 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3301 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     3361 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     3421 tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
     3481 aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct caccgcgcgc
     3541 gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg ggacttcgtg
     3601 gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag cgcggtccag
     3661 gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct ggacgagctg
     3721 tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg gccggccatg
     3781 accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac
     3841 tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt cgattccacc
     3901 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     3961 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     4021 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4081 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     4141 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     4201 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4261 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4321 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4381 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4441 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4501 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4561 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4621 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4681 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4741 ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg
     4801 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4861 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4921 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4981 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     5041 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5101 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5161 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5221 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5281 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5341 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5401 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5461 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5521 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5581 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5641 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5701 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5761 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5821 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5881 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     5941 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     6001 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6061 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6121 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6181 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6241 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6301 acatttcccc gaaaagtgcc acctgacgtc
//
