LOCUS       sequence-348529-        6821 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro transcription of Patiria miniata Dishevelled.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6821)
  AUTHORS   Swartz SZ, Tan TH, Perillo M, Fakhri N, Wessel GM, Wikramanayake AH,
            Cheeseman IM
  TITLE     Polarized Dishevelled dissolution and reassembly drives embryonic 
            axis specification in sea star oocytes.
  JOURNAL   Curr Biol. 2021 Dec 20;31(24):5633-5641.e4. doi: 
            10.1016/j.cub.2021.10.022. Epub 2021 Nov 4.
  PUBMED    34739818
REFERENCE   2  (bases 1 to 6821)
  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"; doi: 
            "10.1016/j.cub.2021.10.022"; journalName: "Curr Biol"; date: 
            "2021-12-20- 20"; volume: "31"; issue: "24"; pages: "5633-5641.e4"
COMMENT     Sequence Label: pZS220
FEATURES             Location/Qualifiers
     source          1..6821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(162..750)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(242..261)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(921..1712,1713..1781))
                     /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     1544..1563
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1782..1886)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1912..2367)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2049..2070)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2261..2280
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2438..3422
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        3457..3475
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3457..3474
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      4785..4794
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5528..6241
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(5558..5586)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     6181..6201
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    6295..6429
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6319..6338)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6373..6392
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(6542..6562)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(6542..6560)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6581..6597)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6581..6597)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6594..6616)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6605..6621
                     /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(6629..6635,6636..6653,6654..6659))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6674..6695
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(join(6812..6821,1..8))
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
       61 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
      121 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
      181 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
      241 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
      301 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
      361 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
      421 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
      481 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
      541 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
      601 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
      661 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
      721 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
      781 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
      841 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
      901 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
      961 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     1021 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     1081 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     1141 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     1201 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     1261 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     1321 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     1381 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     1441 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     1501 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     1561 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     1621 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     1681 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     1741 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     1801 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     1861 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa
     1921 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     1981 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     2041 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     2101 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     2161 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     2221 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     2281 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     2341 cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg
     2401 aagggcgatc ggtgcgggcc tcttcgctat tacgccagtc gaccgccaat tcaatatggc
     2461 gtatatggac tcatgccaat tcaatatggt ggatctggac ctgtgccaat tcaatatggc
     2521 gtatatggac tcgtgccaat tcaatatggt ggatctggac cccagccaat tcaatatggc
     2581 ggacttggca ccatgccaat tcaatatggc ggacctggca ctgtgccaac tggggagggg
     2641 tctacttggc acggtgccaa gtttgaggag gggtcttggc cctgtgccaa gtccgccata
     2701 ttgaattggc atggtgccaa taatggcggc catattggct atatgccagg atcaatatat
     2761 aggcaatatc caatatggcc ctatgccaat atggctattg gccaggttca atactatgta
     2821 ttggccctat gccatatagt attccatata tgggttttcc tattgacgta gatagcccct
     2881 cccaatgggc ggtcccatat accatatatg gggcttccta ataccgccca tagccactcc
     2941 cccattgacg tcaatggtct ctatatatgg tctttcctat tgacgtcata tgggcggtcc
     3001 tattgacgta tatggcgcct cccccattga cgtcaattac ggtaaatggc ccgcctggct
     3061 caatgcccat tgacgtcaat aggaccaccc accattgacg tcaatgggat ggctcattgc
     3121 ccattcatat ccgttctcac gccccctatt gacgtcaatg acggtaaatg gcccacttgg
     3181 cagtacatca atatctatta atagtaactt ggcaagtaca ttactattgg aagtacgcca
     3241 gggtacattg gcagtactcc cattgacgtc aatggcggta aatggcccgc gatggctgcc
     3301 aagtacatcc ccattgacgt caatggggag gggcaatgac gcaaatgggc gttccattga
     3361 cgtaaatggg cggtaggcgt gcctaatggg aggtctatat aagcaatgct cgtttaggga
     3421 accgccattc tgcctgggga cgtcggagca agcttgattt aggtgacact atagaataca
     3481 agctacttgt tctttttgca ggatccacta gtggcgcgcc atggaggaaa cgaagataat
     3541 ataccacata gacgatgagg acaccccgta ccttgtgaag ttgccaattc ctgcggacca
     3601 agtgacatta ggcgatttta agaatgttct caatcggccc aactacaaat tcttcttcgc
     3661 gtctatggac gatgatttcg gcgttgtcaa agaagaaatt gtagaagatg aggccaagct
     3721 gccatgcttc aacggccggg tggtatcctg gctcgtcccg gccgacagca gcacaggagc
     3781 gccatcggac acggggtcgc aatgccaaag cgaagcaccc tcagacatgc agccacccat
     3841 ggaacggacg ggcggcatcg gcgactcgcg accgccttct ttccatgcga atgcaccagg
     3901 aagcacagac aaactcagcg acacagaaac agaatcggtc atcagctcaa gaagagggag
     3961 acaccaaagt tcaaggaggg acagacatca taatgatttt tctcgtccga agagggacca
     4021 ccatcggagc ggtcatgggt acgaaagctc ctccacgtta atgagcagcg acatcgagtc
     4081 tacgagttac ttcgattcca cagacgacga cagcagattt agtagtgcaa cagagcgaag
     4141 cagctcatca cgattaatgc gcagcaaaca caggaaacgg cggcggagac cgcgcatgcc
     4201 tcaagtccag cgtgcctctt cttttagcag tataacagac tctactatgt cactcaacat
     4261 tatcacagta accttaaatt tggacaaaat taacttcttg ggtatcagca ttgttggtca
     4321 aagcaacaag ggcggggacg gtgggatcta cgtgggctcg atcatgaagg ggggtgcggt
     4381 agcagcggac ggcaggatcg agccaggcga catgatcctg caggtcaacg aagtcagctt
     4441 tgagaacatg agcaacgatg atgccgtcag ggtactacga gaagctgtgc accagccagg
     4501 gcccatcaaa cttgtcgttg ctaagtgctg ggatccttca cctaagggtt acttcacaat
     4561 ccctaggagt gagccagtcc ggccgattga tcccggtgcg tgggtggccc acaccaacgc
     4621 catgaaagtg gccgcgtacc acggtcaagc aggaccggtc agtccctcca tgacctcaat
     4681 gacgtccggc agctcttcac taaccagctc tatccccgag tccgaaagat tggaggattt
     4741 cggcccactg tcaatcaact cggacttgaa caccatcgtg cgtgccatgg cggccccgga
     4801 ctctggcctg gacatcagag accgcatgtg gctcaagatt accattccaa acgctttcct
     4861 aggttctgat gttgtggaat ggctgtacaa tcgtgtggag ggtttcacag atcggcgaga
     4921 tgccaggaag tatgcctgca acctcctcaa ggcgggctac atccgacaca ccgtcaacaa
     4981 gatcaccttc tccgagcagt gctactacat ctttggagac ctgtgcggaa acatggcggc
     5041 cctgtcgctg ggtgacgaca ccccagaagt ggaccaggac accctggagc ctctccccca
     5101 gcactggatg ccccccggcc agctaccccc aaatataccc acggcccctc ccctccccta
     5161 ccagcccata ggctattccc agtttgacac ggccagctac accagttttg gaaacgggag
     5221 cgtaggcagt gccggaagtg gaggaagcag tggttcaggc cacagccaag accacaagga
     5281 ccggaaaggc cgcagcaaag gcagctctgg cagcgaatcg gacgccacct ccatcatcag
     5341 ccggtcaacg gcgagagggg acctacccat gcgtgccgac ggctccctgc ggggcgacac
     5401 gccccctatg tccgtcaact ccatgcccat gggcgtggcc atccccccga tgggtcaaat
     5461 gacagggctc gcacaggcca gcgagattgc aagaatgggc cggccagcga tcgccggaca
     5521 tcgattgatg agtaaaggag aagaactttt cactggagtt gtcccaattc ttgttgaatt
     5581 agatggtgat gttaatgggc acaaattttc tgtcagtgga gagggtgaag gtgatgcaac
     5641 atacggaaaa cttaccctta aatttatttg cactactgga aaactacctg ttccatggcc
     5701 aacacttgtc actactctca cttatggtgt tcaatgcttt tcaagatacc cagatcatat
     5761 gaaacggcat gactttttca agagtgccat gcccgaaggt tatgtacagg aaagaactat
     5821 atttttcaaa gatgacggga actacaagac acgtgctgaa gtcaagtttg aaggtgatac
     5881 ccttgttaat agaatcgagt taaaaggtat tgattttaaa gaagatggaa acattcttgg
     5941 acacaaattg gaatacaact ataactcaca caatgtatac atcatggcag acaaacaaaa
     6001 gaatggaatc aaagttaact tcaaaattag acacaacatt gaagatggaa gcgttcaact
     6061 agcagaccat tatcaacaaa atactccaat tggcgatggc cctgtccttt taccagacaa
     6121 ccattacctg tccacacaat ctgccctttc gaaagatccc aacgaaaaga gagaccacat
     6181 ggtccttctt gagtttgtaa cagctgctgg gattacacat ggcatggatg aactatacaa
     6241 ataagaattc aaggcctctc gagcctctag aactatagtg agtcgtatta cgtagatcca
     6301 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     6361 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     6421 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     6481 gaggtttttt aattcgcggc cgcggcgcca atgcattggg cccggtaccc agcttttgtt
     6541 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     6601 gaaattgtta tccgctcaca attccacaca acatacgagc cgggagcata aagtgtaaag
     6661 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     6721 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     6781 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac t
//
