LOCUS       sequence-249855-        4954 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 4954)
  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 4954)
  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_Cerulean_Nanotag_91
FEATURES             Location/Qualifiers
     source          1..4954
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     337..359
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     351..368
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     352..368
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     378..397
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        378..396
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(444..464)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(444..462)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(483..499)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(483..499)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(496..518)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    507..523
                     /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(531..537,538..555,556..561))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    576..597
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        765..910
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          988..1120
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1103..1122
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1174..1183
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1180..1182,1183..1185,1186..1899)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1225..1246)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1486..1505)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1833..1854
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2078..2199
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2089..2108)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2143..2162
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2447..2464)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2618..3206)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2698..2717)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3377..4168,4169..4237))
                     /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     4000..4019
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4238..4342)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4369..4824
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4456..4475)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4666..4687
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(4902..4954,1..487))
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLPWN
                     SSSVPIDREMFWHLHLHWGQPILLVCFVSFCFDGERMLVLSIHTKNQHTDVMKIKIFYC
                     GRSKYLPYLNITYLSW"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctctatcgat aggtaccgag ctcgagttcc
      181 atgggagacg ggcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
      241 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
      301 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
      361 cggccagtga gcgcgcgtaa tacgactcac tatagggcga attgggtacc gggccccccc
      421 tcgaggtcct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
      481 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
      541 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
      601 gcgttgcgct cactgcccgc tttccaccgg tcgtctcagg accgctagcc cgggctcgag
      661 atctgcgatc taagtaagct tgctgcttca tccccgtggc ccgttgctcg cgtttgctgg
      721 cggtgtcccc ggaagaaata tatttgcatg tctttagttc tatgatgaca caaaccccgc
      781 ccagcgtctt gtcattggcg aattcgaaca cgcagatgca gtcggggcgg cgcggtccca
      841 ggtccacttc gcatattaag gtgacgcgtg tggcctcgaa caccgagcga ccctgcagcg
      901 acccgcttaa cagcgtcaac agcgtgccgc agatcccaag cttgcggccg cttaagctgc
      961 agaagttggt cgtgaggcac tgggcaggta agtatcaagg ttacaagaca ggtttaagga
     1021 gaccaataga aactgggctt gtcgagacag agaagactct tgcgtttctg ataggcacct
     1081 attggtctta ctgacatcca ctttgccttt ctctccacag gtgtccactc ccaggttcaa
     1141 tacagctctt aagcggccgc aagctagctt gccgccacca tggtgagcaa gggcgaggag
     1201 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1261 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1321 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctgg
     1381 ggcgtgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1441 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1501 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1561 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caacgccatc
     1621 agcgacaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag
     1681 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1741 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1801 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1861 gccgggatca ctctcggcat ggacgagctg tacaagtgag cctctagact ccctcagcgg
     1921 cgcgcccgga tcctcataag accagtggtg cctaggcagg ctctacggtt ccagtactac
     1981 ccaaccatac gctgacgttt gtgcccgttg taacgcacct gagatcagat ctcacgtgtc
     2041 tagagtcggg gcggccggcc gcttcgagca gacatgataa gatacattga tgagtttgga
     2101 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     2161 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     2221 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac
     2281 aaatgtggta aaatcgataa ggatccgtcg accgatgccc ttgagagcct tcaacccagt
     2341 cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt
     2401 tatcatgcaa ctcgtaggac aggtgccggc agcgctcttc cgcttcctcg ctcactgact
     2461 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     2521 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2581 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2641 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2701 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2761 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct caatgctcac
     2821 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2881 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2941 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     3001 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     3061 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     3121 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     3181 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     3241 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     3301 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     3361 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3421 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3481 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3541 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3601 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3661 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3721 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3781 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3841 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3901 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3961 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     4021 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     4081 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     4141 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     4201 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4261 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4321 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gcgccctgta
     4381 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4441 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4501 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     4561 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     4621 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     4681 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     4741 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     4801 acaaaatatt aacgcttaca atttgccatt cgccattcag gctgcgcaac tgttgggaag
     4861 ggcgatcggt gcgggcctct tcgctattac gccagcccaa gctaccatga taagtaagta
     4921 atattaaggt acgggaggta cttggagcgg ccgc
//
