LOCUS       sequence-258999-        4951 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 4951)
  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 4951)
  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_84
FEATURES             Location/Qualifiers
     source          1..4951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        190..335
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          413..545
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     528..547
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      599..608
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(605..607,608..610,611..1324)
                     /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(650..671)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(911..930)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1258..1279
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1500..1621
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1511..1530)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1565..1584
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1869..1886)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2040..2628)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2120..2139)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2799..3590,3591..3659))
                     /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     3422..3441
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3660..3764)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3791..4246
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3878..3897)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4088..4109
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4324..4863)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLPWN
                     SSSVPIDREMFWHLHLHWGQPILLVCFVSFCFDGERMLVLSIHTKNQHTDVMKIKIFYC
                     GRSKYLPYLNITYLSW"
     polyA_signal    4377..4425
                     /note="synthetic polyadenylation signal"
     misc_feature    4439..4530
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4479..4498
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     4713..4735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4727..4744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4728..4744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4754..4773
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4754..4772
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4820..4840)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4820..4838)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4859..4875)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4859..4875)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4872..4894)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4883..4899
                     /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(4907..4913,4914..4931,4932..4937))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc accggtcgtc
       61 tcaggaccgc tagcccgggc tcgagatctg cgatctaagt aagcttgctg cttcatcccc
      121 gtggcccgtt gctcgcgttt gctggcggtg tccccggaag aaatatattt gcatgtcttt
      181 agttctatga tgacacaaac cccgcccagc gtcttgtcat tggcgaattc gaacacgcag
      241 atgcagtcgg ggcggcgcgg tcccaggtcc acttcgcata ttaaggtgac gcgtgtggcc
      301 tcgaacaccg agcgaccctg cagcgacccg cttaacagcg tcaacagcgt gccgcagatc
      361 ccaagcttgc ggccgcttaa gctgcagaag ttggtcgtga ggcactgggc aggtaagtat
      421 caaggttaca agacaggttt aaggagacca atagaaactg ggcttgtcga gacagagaag
      481 actcttgcgt ttctgatagg cacctattgg tcttactgac atccactttg cctttctctc
      541 cacaggtgtc cactcccagg ttcaatacag ctcttaagcg gccgcaagct agcttgccgc
      601 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      661 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
      721 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
      781 caccctcgtg accaccctga cctggggcgt gcagtgcttc gcccgctacc ccgaccacat
      841 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
      901 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
      961 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1021 gcacaagctg gagtacaacg ccatcagcga caacgtctat atcaccgccg acaagcagaa
     1081 gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1141 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1201 ccactacctg agcacccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat
     1261 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1321 gtgagcctct agactccctc agcggcgcgc cgtaacactg gttttgaagt gcggtcggcg
     1381 tgtaatcctt acgcggatac tgacgtgggt acgtcgcatg gctgatcaca acccgtcagg
     1441 ttaatgagag atctcacgtg tctagagtcg gggcggccgg ccgcttcgag cagacatgat
     1501 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     1561 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     1621 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt
     1681 ttaaagcaag taaaacctct acaaatgtgg taaaatcgat aaggatccgt cgaccgatgc
     1741 ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg actatcgtcg
     1801 ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg gcagcgctct
     1861 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     1921 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     1981 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     2041 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     2101 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     2161 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     2221 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     2281 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     2341 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     2401 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     2461 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     2521 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     2581 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     2641 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     2701 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     2761 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     2821 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     2881 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     2941 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     3001 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     3061 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     3121 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     3181 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     3241 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     3301 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     3361 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     3421 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     3481 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     3541 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     3601 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     3661 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     3721 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     3781 gtgccacctg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3841 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3901 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3961 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     4021 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     4081 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     4141 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     4201 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttgcca ttcgccattc
     4261 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagccc
     4321 aagctaccat gataagtaag taatattaag gtacgggagg tacttggagc ggccgcaata
     4381 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagtactaa
     4441 catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca
     4501 gtgcaagtgc aggtgccaga acatttctct atcgataggt accgagctcg agttccatgg
     4561 gagacgggcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga
     4621 agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc
     4681 aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc
     4741 cagtgagcgc gcgtaatacg actcactata gggcgaattg ggtaccgggc cccccctcga
     4801 ggtcctccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt
     4861 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4921 gaagcataaa gtgtaaagcc tggggtgcct a
//
