LOCUS       sequence-257659-        4938 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 4938)
  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 4938)
  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_105
FEATURES             Location/Qualifiers
     source          1..4938
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(54..593)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLLHN
                     SSSVPIDREMFWHLHLHWGQPILLVCFVSFCFDGERMLVLSIHTKNQHTDVMKIKIFYC
                     GRSKYLPYLNITYLSW"
     polyA_signal    107..155
                     /note="synthetic polyadenylation signal"
     misc_feature    169..260
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     209..228
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     443..465
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     457..474
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     458..474
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     484..503
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        484..502
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(550..570)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(550..568)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(589..605)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(589..605)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(602..624)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    613..629
                     /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(637..643,644..661,662..667))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    682..703
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        871..1016
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          1094..1226
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1209..1228
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1280..1289
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1286..1996
                     /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(1431..1449)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1699..1718
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2168..2289
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2179..2198)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2233..2252
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2537..2554)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2708..3296)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2788..2807)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3467..4258,4259..4327))
                     /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     4090..4109
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4328..4432)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4459..4914
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4546..4565)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4756..4777
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagccc aagctaccat
       61 gataagtaag taatattaag gtacgggagg tacttggagc ggccgcaata aaatatcttt
      121 attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagtactaa catacgctct
      181 ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca gtgcaagtgc
      241 aggtgccaga acatttctct atcgataggt accgagctcg agttgtgcag gagacgggcg
      301 ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga agggcgatcg
      361 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
      421 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc
      481 gcgtaatacg actcactata gggcgaattg ggtaccgggc cccccctcga ggtcctccag
      541 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
      601 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
      661 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
      721 gcccgctttc caccggtcgt ctcacggtgg ctagcccggg ctcgagatct gcgatctaag
      781 taagcttgct gcttcatccc cgtggcccgt tgctcgcgtt tgctggcggt gtccccggaa
      841 gaaatatatt tgcatgtctt tagttctatg atgacacaaa ccccgcccag cgtcttgtca
      901 ttggcgaatt cgaacacgca gatgcagtcg gggcggcgcg gtcccaggtc cacttcgcat
      961 attaaggtga cgcgtgtggc ctcgaacacc gagcgaccct gcagcgaccc gcttaacagc
     1021 gtcaacagcg tgccgcagat cccaagcttg cggccgctta agctgcagaa gttggtcgtg
     1081 aggcactggg caggtaagta tcaaggttac aagacaggtt taaggagacc aatagaaact
     1141 gggcttgtcg agacagagaa gactcttgcg tttctgatag gcacctattg gtcttactga
     1201 catccacttt gcctttctct ccacaggtgt ccactcccag gttcaataca gctcttaagc
     1261 ggccgcaagc tagcttgccg ccaccatggt gagcaagggc gaggaggata acatggccat
     1321 catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt
     1381 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     1441 ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta
     1501 cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt
     1561 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt
     1621 gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac
     1681 caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc
     1741 cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct
     1801 gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt
     1861 gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga
     1921 ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga
     1981 cgagctgtac aagtgagcct ctagactccc tcagcggcgc gcctgagtac accttagtcc
     2041 ggtcgacgag ggagtgtata ccggatcgac gaaagtaatc cgaagagcgt ggctctgttg
     2101 caccgtccat aggaaaagat ctcacgtgtc tagagtcggg gcggccggcc gcttcgagca
     2161 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     2221 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     2281 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     2341 gaggtttttt aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatccgtcg
     2401 accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac
     2461 tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc
     2521 agcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     2581 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     2641 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     2701 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     2761 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2821 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2881 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2941 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     3001 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3061 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     3121 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     3181 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     3241 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     3301 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     3361 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     3421 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     3481 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     3541 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     3601 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     3661 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     3721 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     3781 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3841 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3901 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3961 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     4021 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     4081 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     4141 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     4201 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     4261 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     4321 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     4381 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     4441 cccgaaaagt gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4501 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     4561 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     4621 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     4681 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     4741 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     4801 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     4861 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttgccatt
     4921 cgccattcag gctgcgca
//
