LOCUS       sequence-244933-        5000 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enhancer reporter construct containing the chicken EdnrB enhancer, 
            E2 driving Citrine activity.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5000)
  AUTHORS   Ling ITC, Sauka-Spengler T
  TITLE     Early chromatin shaping predetermines multipotent vagal neural crest
            into neural, neuronal and mesenchymal lineages.
  JOURNAL   Nat Cell Biol. 2019 Dec;21(12):1504-1517. doi: 
            10.1038/s41556-019-0428-9. Epub 2019 Dec 2.
  PUBMED    31792380
REFERENCE   2  (bases 1 to 5000)
  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: 
            "10.1038/s41556-019-0428-9"; journalName: "Nat Cell Biol"; date: 
            "2019-12"; volume: "21"; issue: "12"; pages: "1504-1517"
COMMENT     Sequence Label: pTK-EdnrbE2-Citrine
FEATURES             Location/Qualifiers
     source          1..5000
                     /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"
     promoter        821..966
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          1044..1176
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1159..1178
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1230..1239
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1236..1238,1239..1241,1242..1952)
                     /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(1281..1302)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1542..1561)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1889..1910
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2124..2245
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2135..2154)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2189..2208
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2493..2510)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2664..3252)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2744..2763)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3423..4214,4215..4283))
                     /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     4046..4065
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4284..4388)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4415..4870
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4502..4521)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4712..4733
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctctatcgat aggtaccgag ctcgagttgt
      181 tgcttgtagg accgctcaag tgcttgcagc tcacaattac cctgctggca gttgtaaccg
      241 ccgcactagg actaagtcgc ttgtgtcttg tcattacaat gcttcctttt gtccgatggc
      301 tgcaaatgct tgacagtcat tagaaagtca tccaaggaaa tacttccttt ccatgtacaa
      361 gatgcttgta gtggattttt aacactgaac tttctcagta actcagatag cattagcaaa
      421 agaatagttt cttacagcct gcctctgccc ttaaggttct tttataaaga gaacagaagc
      481 tctttgttgt tactccttca cacgttatta acagaggttt aacagaaaga agaggggctt
      541 gttatagtcc gctgcaattt gtctttttca gttcattcgg atctccatca tgtccaacca
      601 gaagacggat taacccagat ccagggtggc tgcttccata ctaaacagaa aatgaatttt
      661 aaaaagtcag acagtgtaat ctctctctgc caagaaccgg ctagcccggg ctcgagatct
      721 gcgatctaag taagcttgct gcttcatccc cgtggcccgt tgctcgcgtt tgctggcggt
      781 gtccccggaa gaaatatatt tgcatgtctt tagttctatg atgacacaaa ccccgcccag
      841 cgtcttgtca ttggcgaatt cgaacacgca gatgcagtcg gggcggcgcg gtcccaggtc
      901 cacttcgcat attaaggtga cgcgtgtggc ctcgaacacc gagcgaccct gcagcgaccc
      961 gcttaacagc gtcaacagcg tgccgcagat cccaagcttg cggccgctta agctgcagaa
     1021 gttggtcgtg aggcactggg caggtaagta tcaaggttac aagacaggtt taaggagacc
     1081 aatagaaact gggcttgtcg agacagagaa gactcttgcg tttctgatag gcacctattg
     1141 gtcttactga catccacttt gcctttctct ccacaggtgt ccactcccag gttcaataca
     1201 gctcttaagc ggccgcaagc tagcttgccg ccaccatggt gagcaagggc gaggagctgt
     1261 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     1321 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
     1381 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccttc ggctacggcc
     1441 tgatgtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     1501 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     1561 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     1621 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     1681 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     1741 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     1801 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagctaccag tccgccctga
     1861 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     1921 ggatcactct cggcatggac gagctgtaca agtgagcctc tagactccct cagcggcgcg
     1981 cctggtgttc ctcgagatcg gagtcctaaa tcgactgcct tagcggtagg acagtagatc
     2041 agtcagcgac gtgacgaacc ctcgcaggat caagatctca cgtgtctaga gtcggggcgg
     2101 ccggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag
     2161 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     2221 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     2281 tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat
     2341 cgataaggat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc tccttccggt
     2401 gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc atgcaactcg
     2461 taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     2521 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     2581 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2641 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2701 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2761 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2821 tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg taggtatctc
     2881 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2941 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3001 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3061 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     3121 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3181 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3241 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3301 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3361 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3421 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     3481 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     3541 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     3601 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     3661 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     3721 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     3781 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     3841 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     3901 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     3961 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4021 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4081 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4141 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4201 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4261 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4321 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4381 gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg cgcattaagc
     4441 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     4501 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     4561 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     4621 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     4681 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     4741 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     4801 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     4861 cttacaattt gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     4921 gcctcttcgc tattacgcca gcccaagcta ccatgataag taagtaatat taaggtacgg
     4981 gaggtacttg gagcggccgc
//
