LOCUS       sequence-257579-        4526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enhancer cloning nanotag reporter vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4526)
  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 4526)
  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_Citrine_Nanotag_35_neg_Control
FEATURES             Location/Qualifiers
     source          1..4526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        174..319
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          397..529
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     512..531
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      583..592
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(589..591,592..594,595..1305)
                     /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(634..655)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(895..914)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1242..1263
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1484..1605
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1495..1514)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1549..1568
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1853..1870)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2024..2612)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2104..2123)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2783..3574,3575..3643))
                     /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     3406..3425
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3644..3748)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3775..4230
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3862..3881)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4072..4093
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4361..4409
                     /note="synthetic polyadenylation signal"
     misc_feature    4423..4514
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4463..4482
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 cgagctcgag ttgccaggtg ctacgatcga tcggatatca tcgatccagc tggctagccc
       61 gggctcgaga tctgcgatct aagtaagctt gctgcttcat ccccgtggcc cgttgctcgc
      121 gtttgctggc ggtgtccccg gaagaaatat atttgcatgt ctttagttct atgatgacac
      181 aaaccccgcc cagcgtcttg tcattggcga attcgaacac gcagatgcag tcggggcggc
      241 gcggtcccag gtccacttcg catattaagg tgacgcgtgt ggcctcgaac accgagcgac
      301 cctgcagcga cccgcttaac agcgtcaaca gcgtgccgca gatcccaagc ttgcggccgc
      361 ttaagctgca gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag
      421 gtttaaggag accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga
      481 taggcaccta ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc
      541 caggttcaat acagctctta agcggccgca agctagcttg ccgccaccat ggtgagcaag
      601 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
      661 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
      721 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
      781 ttcggctacg gcctgatgtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
      841 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
      901 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
      961 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     1021 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     1081 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     1141 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagctac
     1201 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     1261 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtgagc ctctagactc
     1321 cctcagcggc gcgccgatgc gaaggagttt gcgtaccacg aaccttgctt gagcacaggt
     1381 ccgaggctca actaagggtt tgggacgggc caacgtagct gactgctatg ccagatctca
     1441 cgtgtctaga gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag
     1501 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     1561 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     1621 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     1681 ctctacaaat gtggtaaaat cgataaggat ccgtcgaccg atgcccttga gagccttcaa
     1741 cccagtcagc tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt
     1801 cttctttatc atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca
     1861 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1921 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1981 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     2041 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     2101 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2161 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2221 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2281 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2341 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2401 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2461 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2521 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2581 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2641 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2701 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2761 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     2821 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     2881 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     2941 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     3001 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     3061 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     3121 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     3181 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     3241 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     3301 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     3361 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     3421 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     3481 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     3541 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     3601 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     3661 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     3721 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc
     3781 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3841 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3901 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3961 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     4021 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     4081 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     4141 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     4201 attttaacaa aatattaacg cttacaattt gccattcgcc attcaggctg cgcaactgtt
     4261 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gcccaagcta ccatgataag
     4321 taagtaatat taaggtacgg gaggtacttg gagcggccgc aataaaatat ctttattttc
     4381 attacatctg tgtgttggtt ttttgtgtga atcgatagta ctaacatacg ctctccatca
     4441 aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc cccagtgcaa gtgcaggtgc
     4501 cagaacattt ctctatcgat aggtac
//
