LOCUS       sequence-257649-        4522 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 4522)
  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 4522)
  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_23_neg_Control
FEATURES             Location/Qualifiers
     source          1..4522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..146
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          224..356
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     339..358
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      410..419
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(416..418,419..421,422..1132)
                     /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(461..482)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(722..741)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1069..1090
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1303..1424
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1314..1333)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1368..1387
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1672..1689)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1843..2431)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1923..1942)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2602..3393,3394..3462))
                     /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     3225..3244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3463..3567)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3594..4049
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3681..3700)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3891..3912
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4180..4228
                     /note="synthetic polyadenylation signal"
     misc_feature    4242..4333
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4282..4301
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 atgacacaaa ccccgcccag cgtcttgtca ttggcgaatt cgaacacgca gatgcagtcg
       61 gggcggcgcg gtcccaggtc cacttcgcat attaaggtga cgcgtgtggc ctcgaacacc
      121 gagcgaccct gcagcgaccc gcttaacagc gtcaacagcg tgccgcagat cccaagcttg
      181 cggccgctta agctgcagaa gttggtcgtg aggcactggg caggtaagta tcaaggttac
      241 aagacaggtt taaggagacc aatagaaact gggcttgtcg agacagagaa gactcttgcg
      301 tttctgatag gcacctattg gtcttactga catccacttt gcctttctct ccacaggtgt
      361 ccactcccag gttcaataca gctcttaagc ggccgcaagc tagcttgccg ccaccatggt
      421 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
      481 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
      541 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
      601 gaccaccttc ggctacggcc tgatgtgctt cgcccgctac cccgaccaca tgaagcagca
      661 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
      721 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
      781 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
      841 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
      901 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
      961 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1021 gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1081 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtgagcctc
     1141 tagactccct cagcggcgcg ccgcgagatt agtcgcccag aatcaggcga gcttaccggt
     1201 ttccaccatt atacccacga catgacaggc ggggatttcc ggctctcgta gagatctcac
     1261 gtgtctagag tcggggcggc cggccgcttc gagcagacat gataagatac attgatgagt
     1321 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     1381 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     1441 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     1501 tctacaaatg tggtaaaatc gataaggatc cgtcgaccga tgcccttgag agccttcaac
     1561 ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact tatgactgtc
     1621 ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tcttccgctt cctcgctcac
     1681 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     1741 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     1801 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     1861 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     1921 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     1981 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2041 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2101 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     2161 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     2221 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     2281 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     2341 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     2401 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     2461 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     2521 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     2581 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     2641 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     2701 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     2761 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     2821 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     2881 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     2941 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3001 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     3061 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     3121 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     3181 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     3241 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     3301 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     3361 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     3421 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     3481 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     3541 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc
     3601 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     3661 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     3721 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     3781 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     3841 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     3901 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     3961 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     4021 ttttaacaaa atattaacgc ttacaatttg ccattcgcca ttcaggctgc gcaactgttg
     4081 ggaagggcga tcggtgcggg cctcttcgct attacgccag cccaagctac catgataagt
     4141 aagtaatatt aaggtacggg aggtacttgg agcggccgca ataaaatatc tttattttca
     4201 ttacatctgt gtgttggttt tttgtgtgaa tcgatagtac taacatacgc tctccatcaa
     4261 aacaaaacga aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag tgcaggtgcc
     4321 agaacatttc tctatcgata ggtaccgagc tcgagttgcc agagctggat cgatgatatc
     4381 cgatcgatcg tagcactgtt ggctagcccg ggctcgagat ctgcgatcta agtaagcttg
     4441 ctgcttcatc cccgtggccc gttgctcgcg tttgctggcg gtgtccccgg aagaaatata
     4501 tttgcatgtc tttagttcta tg
//
