LOCUS       sequence-258625-        5539 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  enhancer reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5539)
  AUTHORS   Williams RM, Candido-Ferreira I, Repapi E, Gavriouchkina D, 
            Senanayake U, Ling ITC, Telenius J, Taylor S, Hughes J, 
            Sauka-Spengler T
  TITLE     Reconstruction of the Global Neural Crest Gene Regulatory Network In
            Vivo.
  JOURNAL   Dev Cell. 2019 Oct 21;51(2):255-276.e7. doi: 
            10.1016/j.devcel.2019.10.003.
  PUBMED    31639368
REFERENCE   2  (bases 1 to 5539)
  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.devcel.2019.10"; journalName: "Dev Cell"; date: 
            "2019-10-21- 21"; volume: "51"; issue: "2"; pages: "255-276.e7"
COMMENT     Sequence Label: pTK_Brn3_enh_250_Citrine
FEATURES             Location/Qualifiers
     source          1..5539
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    14..105
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     54..73
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        1311..1456
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          1534..1666
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1649..1668
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1720..1729
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1726..1728,1729..1731,1732..2442)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1771..1792)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2032..2051)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2379..2400
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2614..2735
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2625..2644)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2679..2698
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2983..3000)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3154..3742)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3234..3253)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3913..4704,4705..4773))
                     /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     4536..4555
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4774..4878)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4905..5360
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4992..5011)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5202..5223
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    5491..5539
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 aatcgatagt actaacatac gctctccatc aaaacaaaac gaaacaaaac aaactagcaa
       61 aataggctgt ccccagtgca agtgcaggtg ccagaacatt tctctatcga taggtaccga
      121 gctcgagttg ccaggttcca gtgcacaaga aattcgtaga atcatagaat gtttgggttg
      181 gaagagatct taatcccatg taattacaac tccttgccat gggctggctg cctccaccag
      241 ctcaggctgc ccagggcccc atccaacctg gggcacctcc agtgatgggg cacccacagt
      301 tccctgggca gcagtgccag ggcctcacca attcaccatt tctggctcaa ctcctgctaa
      361 agctctggaa aggaccatga ggaaaaggca gtgggtatac cccattggct tccgaccaac
      421 acacccacct ttatcatccc accattggcc aaagtgtccg agcttgcaga gtgagccgag
      481 acagctgatg agaaggatgt ggcagcggtc acctcggggc agctgtaggg cctctgatcc
      541 atcattctgc agaaagtcct ccgtgtgtct tttcccctgg atcctctcct ggaggcctgg
      601 aaagctgtgt cccctctgcc ctgtgttcct attaatcgct gccagtgggt ataattggta
      661 gctagagcac acagacccca ttaaagccca tgagggtgcc aaatgctctc attcaggcac
      721 aaatgaagga gcaatcaaac ccacttaggt aataaaaaca gctggagtct gcaggcaacc
      781 gcccgaggca ccaaccacat ggcaggcagg acaatcctct gaaaagtctg cattgaggtg
      841 tcctttgatt gaaaacgtgg ggattaggag gagcacagca aggtttcatt ggctaaaggc
      901 agacagacag aaggagaaac tatgagttgt cttccaaaaa caacagggag aaggagtttc
      961 ttaggcacag agtttgccgg agacaattgg ttccctatgg gaatgatgag cgagagcatc
     1021 tgtaggtgtc atggtaccac ggtgtcccca actccttttg ctaacttcgc atgccgtggc
     1081 agataaccac cacatctgag cctcacaagg agcagcaatg acccgaagag ctttctggct
     1141 catcacacag atgtgatgtg taggggattg tctcaggatg aatctgttgg ctagcccggg
     1201 ctcgagatct gcgatctaag taagcttgct gcttcatccc cgtggcccgt tgctcgcgtt
     1261 tgctggcggt gtccccggaa gaaatatatt tgcatgtctt tagttctatg atgacacaaa
     1321 ccccgcccag cgtcttgtca ttggcgaatt cgaacacgca gatgcagtcg gggcggcgcg
     1381 gtcccaggtc cacttcgcat attaaggtga cgcgtgtggc ctcgaacacc gagcgaccct
     1441 gcagcgaccc gcttaacagc gtcaacagcg tgccgcagat cccaagcttg cggccgctta
     1501 agctgcagaa gttggtcgtg aggcactggg caggtaagta tcaaggttac aagacaggtt
     1561 taaggagacc aatagaaact gggcttgtcg agacagagaa gactcttgcg tttctgatag
     1621 gcacctattg gtcttactga catccacttt gcctttctct ccacaggtgt ccactcccag
     1681 gttcaataca gctcttaagc ggccgcaagc tagcttgccg ccaccatggt gagcaagggc
     1741 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     1801 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     1861 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccttc
     1921 ggctacggcc tgatgtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc
     1981 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     2041 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     2101 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     2161 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     2221 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     2281 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagctaccag
     2341 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     2401 accgccgccg ggatcactct cggcatggac gagctgtaca agtgagcctc tagactccct
     2461 cagcggcgcg cctggtgttc ctcgagatcg gagtcctaaa tcgactgcct tagcggtagg
     2521 acagtagatc agtcagcgac gtgacgaacc ctcgcaggat caagatccca cgtgtctaga
     2581 gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     2641 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     2701 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     2761 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     2821 gtggtaaaat cgataaggat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc
     2881 tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc
     2941 atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct
     3001 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3061 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3121 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3181 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3241 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3301 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3361 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3421 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3481 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3541 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3601 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3661 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3721 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3781 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3841 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3901 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3961 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4021 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4081 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4141 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4201 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4261 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4321 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4381 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4441 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4501 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4561 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4621 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4681 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4741 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4801 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4861 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg
     4921 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     4981 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     5041 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     5101 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     5161 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     5221 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     5281 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     5341 aatattaacg cttacaattt gccattcgcc attcaggctg cgcaactgtt gggaagggcg
     5401 atcggtgcgg gcctcttcgc tattacgcca gcccaagcta ccatgataag taagtaatat
     5461 taaggtacgg gaggtacttg gagcggccgc aataaaatat ctttattttc attacatctg
     5521 tgtgttggtt ttttgtgtg
//
