LOCUS       sequence-257643-        4942 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 4942)
  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 4942)
  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_BsmBI_LacZ_Cherry_Nanotag_106
FEATURES             Location/Qualifiers
     source          1..4942
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        63..208
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          286..418
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     401..420
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      472..481
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             478..1188
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(623..641)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     891..910
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1364..1485
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1375..1394)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1429..1448
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1733..1750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1904..2492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1984..2003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2663..3454,3455..3523))
                     /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     3286..3305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3524..3628)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3655..4110
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3742..3761)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3952..3973
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4188..4727)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLLHN
                     SSSVPIDREMFWHLHLHWGQPILLVCFVSFCFDGERMLVLSIHTKNQHTDVMKIKIFYC
                     GRSKYLPYLNITYLSW"
     polyA_signal    4241..4289
                     /note="synthetic polyadenylation signal"
     misc_feature    4303..4394
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4343..4362
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     4577..4599
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4591..4608
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4592..4608
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4618..4637
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4618..4636
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4684..4704)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4684..4702)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4723..4739)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4723..4739)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4736..4758)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4747..4763
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4771..4777,4778..4795,4796..4801))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4816..4837
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gttgctcgcg tttgctggcg gtgtccccgg aagaaatata tttgcatgtc tttagttcta
       61 tgatgacaca aaccccgccc agcgtcttgt cattggcgaa ttcgaacacg cagatgcagt
      121 cggggcggcg cggtcccagg tccacttcgc atattaaggt gacgcgtgtg gcctcgaaca
      181 ccgagcgacc ctgcagcgac ccgcttaaca gcgtcaacag cgtgccgcag atcccaagct
      241 tgcggccgct taagctgcag aagttggtcg tgaggcactg ggcaggtaag tatcaaggtt
      301 acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg
      361 cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt
      421 gtccactccc aggttcaata cagctcttaa gcggccgcaa gctagcttgc cgccaccatg
      481 gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg
      541 cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
      601 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
      661 gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc
      721 gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
      781 atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc
      841 gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg
      901 cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc
      961 ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag
     1021 gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac
     1081 atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc
     1141 gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtgagc ctctagactc
     1201 cctcagcggc gcgccgctgc atctcagact tacgccgata cgtttctcag tcggccgctc
     1261 ttaggctagt taggtagaac gccgaagctc ccaaagctta caccgttgag taagatctca
     1321 cgtgtctaga gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag
     1381 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     1441 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     1501 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     1561 ctctacaaat gtggtaaaat cgataaggat ccgtcgaccg atgcccttga gagccttcaa
     1621 cccagtcagc tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt
     1681 cttctttatc atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca
     1741 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     1921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     2701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     2761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     2821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     2881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     2941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     3001 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     3061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     3121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     3181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     3241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     3301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     3361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     3421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     3481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     3541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     3601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc
     3661 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3721 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3781 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3841 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     3901 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     3961 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     4021 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     4081 attttaacaa aatattaacg cttacaattt gccattcgcc attcaggctg cgcaactgtt
     4141 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gcccaagcta ccatgataag
     4201 taagtaatat taaggtacgg gaggtacttg gagcggccgc aataaaatat ctttattttc
     4261 attacatctg tgtgttggtt ttttgtgtga atcgatagta ctaacatacg ctctccatca
     4321 aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc cccagtgcaa gtgcaggtgc
     4381 cagaacattt ctctatcgat aggtaccgag ctcgagttgt gcaggagacg ggcgccgcta
     4441 cagggcgcgt cccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     4501 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
     4561 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga gcgcgcgtaa
     4621 tacgactcac tatagggcga attgggtacc gggccccccc tcgaggtcct ccagcttttg
     4681 ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca tggtcatagc tgtttcctgt
     4741 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4801 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4861 tttccaccgg tcgtctcacg gtggctagcc cgggctcgag atctgcgatc taagtaagct
     4921 tgctgcttca tccccgtggc cc
//
