LOCUS       sequence-258504-        7016 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 7016)
  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 7016)
  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_TFAP2b_enh_32_Citrine
FEATURES             Location/Qualifiers
     source          1..7016
                     /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"
     CDS             complement(1056..1067)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     promoter        2786..2931
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          3009..3141
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3124..3143
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      3195..3204
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3201..3203,3204..3206,3207..3917)
                     /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(3246..3267)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3507..3526)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3854..3875
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4091..4212
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4102..4121)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4156..4175
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4460..4477)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4631..5219)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4711..4730)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5390..6181,6182..6250))
                     /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     6013..6032
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6251..6355)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6382..6837
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6469..6488)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6679..6700
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    6968..7016
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 aatcgatagt actaacatac gctctccatc aaaacaaaac gaaacaaaac aaactagcaa
       61 aataggctgt ccccagtgca agtgcaggtg ccagaacatt tctctatcga taggtaccga
      121 gctcgagttg ccaggtcacg cgaataaaac agcacaatgt agcaatagac ctatgttaat
      181 cacacttctt ctgaaaacag caaaaaaaaa gcttcttttt cagcatcttg ttaatatcac
      241 tgtatgatac aaaatgaacc ttgaaactca gagttaaatt gaatttggat acattttatg
      301 gaagacctag cgaattttac tttttttcct gatttacgaa ctgatttgta aaatctataa
      361 cagatgctga ctgcttccat ttttgattgc ttttatttta tctttctaca ctcaggagaa
      421 gcttatgaat ccagactgat gcaaaggtaa ctgctaccct tcaatattcg accctatctt
      481 ctatcagaag actcttacta attgttggag actcatttac cagcaaaggg taagaagtca
      541 aacctccatt accttgctgg aaaatgccaa taaccaacag tcctggaatg tcctcttcac
      601 atggtgtggg ttagatggtc agctggatta aatttaacat cactgatgga gagaagcagt
      661 gcatcaaatg caaatggaac agagagaaca agcagctact cttttacctg tcttatggac
      721 tcctggctac acagccctta tctccccata cacacaaagg cctgaagtag ccatgttgct
      781 tgagttggac atacctccca gtggtaatgt ataatttttc ttgtcttagt tatgtgaata
      841 acaacaaaga caggcttttt tttttcttcc caattttatg ttgctttcaa tctgacatca
      901 gcgttccatt actgagttaa ctggcgtcca cgtggctttg taactaccaa cagcagaagt
      961 ggaaatcaaa ccatatggca gggctgtgct tcagaaagtt taagagtctg ccatttagct
     1021 tggagatttt tagttccttt tgattcagac tccaattgga gttcccataa atcctgcaaa
     1081 tagaacaatt tctttttttc tcatttaaat cccatatact gccttttcat aaaaatttct
     1141 tctataaaag aggaaaagta taattctact ttttccatga gagggaatgg acatcctcta
     1201 cttttagcaa ttatttatca caaagaaaga aaactcacta aaatagaaaa gatcagcctt
     1261 taaaaaataa gaaagttgtc aacactttgg ccatcacaaa ctcagggaga gaacttatct
     1321 ctcatacgtt cactgaaatc agtttgttaa tgaagaaaca aggtaaagaa tgggaaatag
     1381 aaacattgtg ctttctaggc ctttattttg taaacatttg gacttgtgca aattcgttaa
     1441 ttgaggcaat tgatttttgc ctgaaaatgg tagcaatgtt taattcatta aagcctaaaa
     1501 taacatcctt tgggctaaat ggggtcattt catttgtaac cttttagtga tgtattaaca
     1561 ttttggaagc atttctgtgc atatcttaaa aagtgccaca ttaacaggaa cgggcctgga
     1621 gaaagaaaag aagttgcaaa cactcacaca cgcacacagg cacacacaca caatatatat
     1681 ggcctccagc ccctgtcctt cctcttctgc agcattttaa ggtctgtgtt taatattgcc
     1741 aagcctactt gagtgataaa tgcaactgta taaaaattac ttattacaag ctgtcatgtt
     1801 gcctcactca cattgcaggg ctagaggcac tgtacataaa gctcgaatct ccaccttgac
     1861 agcatcggca gggctggctt taaaataaaa tgctttcagt catgaataca aaacataatt
     1921 taccttttca taatacttca gtttggaaac actttggaaa gctcatgtta ccatcaaacc
     1981 tttacctgag ccttgccagc acccagttaa ttccgagttg tcaatttact tcagtgccgc
     2041 tttccatcac tcagcctttg aagaaacatt aagctgttca aattgccatc cctttaattt
     2101 tttttttatt ttttttttta agttatattt aacctcgatt ctttggcagc gagtagttga
     2161 cacaatagga ttatcccaag cattaaatta caatttggga gagtgcctgc tcagcgcact
     2221 tcttttgcag aagccccttg acttgctgtg tgttcccagc accaaaaatt atcctgtctt
     2281 ccatccccct gttttccccc tctcttttgc atagcgccca gttacacaat cccccttgag
     2341 ttgctcctcc ctcccagaga ggttcagagg tttaattctg tgccgtggtt gcccagccac
     2401 cccttttctc ctgcattagt actgtattga tgcttaaata taaaagctca ctgtaagctg
     2461 tggatgtcag tgagattgaa attgaaagtg aaggttctgg ggaggctggc tgggtaacaa
     2521 ctgatgcgtt gtgactgttt ttatagttgg tcatgatgca tacagtatgt gaagttccca
     2581 tctgtactgg tgtctgccca ccagccaccc tgccacattc gcaaaagtgg ctccagcacc
     2641 gagcgcaata ggaacagact gttggctagc ccgggctcga gatctgcgat ctaagtaagc
     2701 ttgctgcttc atccccgtgg cccgttgctc gcgtttgctg gcggtgtccc cggaagaaat
     2761 atatttgcat gtctttagtt ctatgatgac acaaaccccg cccagcgtct tgtcattggc
     2821 gaattcgaac acgcagatgc agtcggggcg gcgcggtccc aggtccactt cgcatattaa
     2881 ggtgacgcgt gtggcctcga acaccgagcg accctgcagc gacccgctta acagcgtcaa
     2941 cagcgtgccg cagatcccaa gcttgcggcc gcttaagctg cagaagttgg tcgtgaggca
     3001 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
     3061 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
     3121 actttgcctt tctctccaca ggtgtccact cccaggttca atacagctct taagcggccg
     3181 caagctagct tgccgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     3241 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     3301 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     3361 tgcccgtgcc ctggcccacc ctcgtgacca ccttcggcta cggcctgatg tgcttcgccc
     3421 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     3481 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     3541 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     3601 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     3661 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     3721 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     3781 tgctgctgcc cgacaaccac tacctgagct accagtccgc cctgagcaaa gaccccaacg
     3841 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     3901 tggacgagct gtacaagtga gcctctagac tccctcagcg gcgcgccgtt gcgttccaaa
     3961 cgtcgtggag ctccctgtca agacacgaac cgctaacgcc accttagtcc gtgaagcatg
     4021 atacgcgaca taccccagga gatctcacgt gtctagagtc ggggcggccg gccgcttcga
     4081 gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa
     4141 aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc
     4201 aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg
     4261 tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtaaaatcga taaggatccg
     4321 tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat
     4381 gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc
     4441 ggcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4501 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4561 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4621 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     4681 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     4741 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     4801 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     4861 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     4921 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4981 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5041 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5101 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5161 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5221 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5281 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5341 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5401 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5461 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     5521 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     5581 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     5641 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     5701 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     5761 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     5821 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     5881 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     5941 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6001 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6061 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6121 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6181 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6241 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6301 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6361 ttccccgaaa agtgccacct gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     6421 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     6481 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     6541 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     6601 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     6661 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     6721 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     6781 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttgcc
     6841 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     6901 tacgccagcc caagctacca tgataagtaa gtaatattaa ggtacgggag gtacttggag
     6961 cggccgcaat aaaatatctt tattttcatt acatctgtgt gttggttttt tgtgtg
//
