LOCUS       sequence-257646-        4525 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 4525)
  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 4525)
  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_5_neg_Control
FEATURES             Location/Qualifiers
     source          1..4525
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        60..205
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          283..415
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     398..417
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      469..478
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(475..477,478..480,481..1191)
                     /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(520..541)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(781..800)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1128..1149
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1365..1486
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1376..1395)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1430..1449
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1734..1751)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1905..2493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1985..2004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2664..3455,3456..3524))
                     /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     3287..3306
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3525..3629)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3656..4111
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3743..3762)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3953..3974
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4242..4290
                     /note="synthetic polyadenylation signal"
     misc_feature    4304..4395
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4344..4363
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 gctcgcgttt gctggcggtg tccccggaag aaatatattt gcatgtcttt agttctatga
       61 tgacacaaac cccgcccagc gtcttgtcat tggcgaattc gaacacgcag atgcagtcgg
      121 ggcggcgcgg tcccaggtcc acttcgcata ttaaggtgac gcgtgtggcc tcgaacaccg
      181 agcgaccctg cagcgacccg cttaacagcg tcaacagcgt gccgcagatc ccaagcttgc
      241 ggccgcttaa gctgcagaag ttggtcgtga ggcactgggc aggtaagtat caaggttaca
      301 agacaggttt aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt
      361 ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc cacaggtgtc
      421 cactcccagg ttcaatacag ctcttaagcg gccgcaagct agcttgccgc caccatggtg
      481 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
      541 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
      601 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
      661 accaccttcg gctacggcct gatgtgcttc gcccgctacc ccgaccacat gaagcagcac
      721 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
      781 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
      841 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
      901 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
      961 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     1021 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1081 agctaccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1141 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtgagcctct
     1201 agactccctc agcggcgcgc cgttgcgttc caaacgtcgt ggagctccct gtcaagacac
     1261 gaaccgctaa cgccacctta gtccgtgaag catgatacgc gacatacccc aggagatctc
     1321 acgtgtctag agtcggggcg gccggccgct tcgagcagac atgataagat acattgatga
     1381 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga
     1441 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg
     1501 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa
     1561 cctctacaaa tgtggtaaaa tcgataagga tccgtcgacc gatgcccttg agagccttca
     1621 acccagtcag ctccttccgg tgggcgcggg gcatgactat cgtcgccgca cttatgactg
     1681 tcttctttat catgcaactc gtaggacagg tgccggcagc gctcttccgc ttcctcgctc
     1741 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     1801 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     1861 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     1921 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     1981 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2041 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2101 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2161 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2221 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2281 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2341 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2401 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2461 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2521 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     2581 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     2641 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     2701 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     2761 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     2821 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     2881 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     2941 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3001 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3061 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3121 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3181 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3241 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3301 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3361 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3421 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3481 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3541 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     3601 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgcg
     3661 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     3721 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     3781 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     3841 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     3901 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     3961 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     4021 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     4081 aattttaaca aaatattaac gcttacaatt tgccattcgc cattcaggct gcgcaactgt
     4141 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agcccaagct accatgataa
     4201 gtaagtaata ttaaggtacg ggaggtactt ggagcggccg caataaaata tctttatttt
     4261 cattacatct gtgtgttggt tttttgtgtg aatcgatagt actaacatac gctctccatc
     4321 aaaacaaaac gaaacaaaac aaactagcaa aataggctgt ccccagtgca agtgcaggtg
     4381 ccagaacatt tctctatcga taggtaccga gctcgagttg ccagagctgg atcgatgata
     4441 tccgatcgat cgtagcactg ttggctagcc cgggctcgag atctgcgatc taagtaagct
     4501 tgctgcttca tccccgtggc ccgtt
//
