LOCUS       sequence-137870-        4719 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 4719)
  AUTHORS   Dorsky RI, Raible DW, Moon RT
  TITLE     Direct regulation of nacre, a zebrafish MITF homolog required for 
            pigment cell formation, by the Wnt pathway.
  JOURNAL   Genes Dev. 2000 Jan 15;14(2):158-62.
  PUBMED    10652270
REFERENCE   2  (bases 1 to 4719)
  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."; date: "2000-01-15"; volume: "14(2)"; pages: "158-62"
COMMENT     Sequence Label: M28 Nac>GFP
FEATURES             Location/Qualifiers
     source          1..4719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      895..904
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             912..1625
                     /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(942..970)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1565..1585
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    1689..1810
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1700..1719)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1754..1773
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2058..2075)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2229..2817)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2309..2328)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2988..3779,3780..3848))
                     /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     3611..3630
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3849..3953)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3980..4435
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4067..4086)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4277..4298
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4566..4614
                     /note="synthetic polyadenylation signal"
     misc_feature    4628..4719
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4668..4687
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 atcgataggt accgagctct tacgcgtgct agcttgttta tgattgttag cttttttaac
       61 tacagttcat ttctatgctt tctgtgcaaa tgcacaacag ttactgattt actctgataa
      121 taattgtgcc agtacatcca acaacaacaa aaaaaaaaga cttacaatca agccccaaag
      181 tgttcatacg tctggcaaat tctgacttaa agttactttt attcaaccag ctagtttttt
      241 tttgactaaa tgacacgtaa gataataaga tgatgtacat gaggcatcat tgtagaaaaa
      301 aaaaactctt tctcaggttt tatttattta agtcaaaatt tgggttatga atcatttcgg
      361 gcttgactgt acatgtctga atccattggc ttctgaatat gaactgtacg tctgtgattt
      421 tgtttgtgcg cacttacata cgtgtgcaca atatttcaac tcttaagaaa aaaatgcaat
      481 taccaaaggc ccatcagaca acaaagttct ctttcctatc ttttgctctt caacatttca
      541 gagataactc catcttaaac tgtcttgttt ttagaccaac agtgctagtg attgtatgcc
      601 ggatgctgag aacaaacaat gttttatgca ggatgagtgg aactaaagat gaaacgttag
      661 ttagccgtaa gccagtttca gtttagacac tagtaaccca tcgtctcaaa gaggcttttg
      721 tcgaatcgga ccggtgacgt caaatgaact ttataaaggc ataaattcat cgtttgggta
      781 aaaaaaacaa tatgagaaga gcaatcccct cccaccccct taaatacatt ttgtgtcttt
      841 tctttggcat tgacatcttc agctggagct tggcattccg gtactgttgg taaagccacc
      901 atggatccac catgagtaaa ggagaagaac ttttcactgg agttgtccca attcttgttg
      961 aattagatgg tgatgttaat gggcacaaat tttctgtcag tggagagggt gaaggtgatg
     1021 caacatacgg aaaacttacc cttaaattta tttgcactac tggaaaacta cctgttccat
     1081 ggccaacact tgtcactact ctcacttatg gtgttcaatg cttttcaaga tacccagatc
     1141 atatgaaacg gcatgacttt ttcaagagtg ccatgcccga aggttatgta caggaaagaa
     1201 ctatattttt caaagatgac gggaactaca agacacgtgc tgaagtcaag tttgaaggtg
     1261 atacccttgt taatagaatc gagttaaaag gtattgattt taaagaagat ggaaacattc
     1321 ttggacacaa attggaatac aactataact cacacaatgt atacatcatg gcagacaaac
     1381 aaaagaatgg aatcaaagtt aacttcaaaa ttagacacaa cattgaagat ggaagcgttc
     1441 aactagcaga ccattatcaa caaaatactc caattggcga tggccctgtc cttttaccag
     1501 acaaccatta cctgtccaca caatctgccc tttcgaaaga tcccaacgaa aagagagacc
     1561 acatggtcct tcttgagttt gtaacagctg ctgggattac acatggcatg gatgaactat
     1621 acaaataaga attcaaggcc tctcgagcct ctagagtcgg ggcggccggc cgcttcgagc
     1681 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1741 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     1801 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     1861 ggaggttttt taaagcaagt aaaacctcta caaatgtggt aaaatcgata aggatccgtc
     1921 gaccgatgcc cttgagagcc ttcaacccag tcagctcctt ccggtgggcg cggggcatga
     1981 ctatcgtcgc cgcacttatg actgtcttct ttatcatgca actcgtagga caggtgccgg
     2041 cagcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     2101 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2161 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2221 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2281 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2341 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2401 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2461 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     2521 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     2581 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     2641 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     2701 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     2761 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     2821 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     2881 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     2941 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3001 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     3061 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     3121 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     3181 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3241 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3301 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3361 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3421 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3481 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     3541 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     3601 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     3661 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     3721 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     3781 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     3841 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     3901 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     3961 ccccgaaaag tgccacctga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4021 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4081 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4141 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4201 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4261 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4321 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     4381 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttgccat
     4441 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     4501 cgccagccca agctaccatg ataagtaagt aatattaagg tacgggaggt acttggagcg
     4561 gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga
     4621 tagtactaac atacgctctc catcaaaaca aaacgaaaca aaacaaacta gcaaaatagg
     4681 ctgtccccag tgcaagtgca ggtgccagaa catttctct
//
