LOCUS       sequence-257595-        4528 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 4528)
  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 4528)
  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_32_neg_Control
FEATURES             Location/Qualifiers
     source          1..4528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        93..238
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          316..448
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     431..450
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      502..511
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(508..510,511..513,514..1224)
                     /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(553..574)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(814..833)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1161..1182
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1401..1522
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1412..1431)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1466..1485
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1770..1787)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1941..2529)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2021..2040)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2700..3491,3492..3560))
                     /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     3323..3342
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3561..3665)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3692..4147
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3779..3798)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3989..4010
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4278..4326
                     /note="synthetic polyadenylation signal"
     misc_feature    4340..4431
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4380..4399
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 agtaagcttg ctgcttcatc cccgtggccc gttgctcgcg tttgctggcg gtgtccccgg
       61 aagaaatata tttgcatgtc tttagttcta tgatgacaca aaccccgccc agcgtcttgt
      121 cattggcgaa ttcgaacacg cagatgcagt cggggcggcg cggtcccagg tccacttcgc
      181 atattaaggt gacgcgtgtg gcctcgaaca ccgagcgacc ctgcagcgac ccgcttaaca
      241 gcgtcaacag cgtgccgcag atcccaagct tgcggccgct taagctgcag aagttggtcg
      301 tgaggcactg ggcaggtaag tatcaaggtt acaagacagg tttaaggaga ccaatagaaa
      361 ctgggcttgt cgagacagag aagactcttg cgtttctgat aggcacctat tggtcttact
      421 gacatccact ttgcctttct ctccacaggt gtccactccc aggttcaata cagctcttaa
      481 gcggccgcaa gctagcttgc cgccaccatg gtgagcaagg gcgaggagct gttcaccggg
      541 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
      601 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
      661 ggcaagctgc ccgtgccctg gcccaccctc gtgaccacct tcggctacgg cctgatgtgc
      721 ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
      781 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
      841 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
      901 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
      961 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     1021 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1081 ggccccgtgc tgctgcccga caaccactac ctgagctacc agtccgccct gagcaaagac
     1141 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1201 ctcggcatgg acgagctgta caagtgagcc tctagactcc ctcagcggcg cgccgctctc
     1261 cagaaacttc gcaaacacct atgcctaacg ccagccgaat cctgcagtcc gttacctagt
     1321 cgaaaggcat tctgggagca tcgcgtcaca gatctcacgt gtctagagtc ggggcggccg
     1381 gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     1441 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     1501 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     1561 gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtaaaatcga
     1621 taaggatccg tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg
     1681 cgcggggcat gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag
     1741 gacaggtgcc ggcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     1801 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     1861 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1921 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     1981 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     2041 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     2101 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     2161 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     2221 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     2281 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     2341 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     2401 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     2461 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     2521 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     2581 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     2641 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     2701 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     2761 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     2821 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     2881 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     2941 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     3001 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     3061 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     3121 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     3181 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     3241 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     3301 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     3361 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     3421 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     3481 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     3541 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     3601 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     3661 ccgcgcacat ttccccgaaa agtgccacct gacgcgccct gtagcggcgc attaagcgcg
     3721 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3781 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3841 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3901 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3961 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4021 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4081 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     4141 acaatttgcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     4201 tcttcgctat tacgccagcc caagctacca tgataagtaa gtaatattaa ggtacgggag
     4261 gtacttggag cggccgcaat aaaatatctt tattttcatt acatctgtgt gttggttttt
     4321 tgtgtgaatc gatagtacta acatacgctc tccatcaaaa caaaacgaaa caaaacaaac
     4381 tagcaaaata ggctgtcccc agtgcaagtg caggtgccag aacatttctc tatcgatagg
     4441 taccgagctc gagttgccag agctggatcg atgatatccg atcgatcgta gcactgttgg
     4501 ctagcccggg ctcgagatct gcgatcta
//
