LOCUS       sequence-317737-        5228 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector based on the LiOn integration-coupled translational switch 
            (Kumamoto et al bioRxiv 2019) expressing the fluorescent protein 
            EGFP from a CAG promoter upon action of the piggyBac transposase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5228)
  AUTHORS   Kumamoto T, Maurinot F, Barry-Martinet R, Vaslin C, 
            Vandormael-Pournin S, Le M, Lerat M, Niculescu D, Cohen-Tannoudji M,
            Rebsam A, Loulier K, Nedelec S, Tozer S, Livet J
  TITLE     Direct Readout of Neural Stem Cell Transgenesis with an 
            Integration-Coupled Gene Expression Switch.
  JOURNAL   Neuron. 2020 Aug 19;107(4):617-630.e6. doi: 
            10.1016/j.neuron.2020.05.038. Epub 2020 Jun 18.
  PUBMED    32559415
REFERENCE   2  (bases 1 to 5228)
  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.neuron.2020.05.038"; journalName: "Neuron"; date: 
            "2020-08-19- 19"; volume: "107"; issue: "4"; pages: "617-630.e6"
COMMENT     Sequence Label: LiOn-CAG∞GFP
FEATURES             Location/Qualifiers
     source          1..5228
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..657
                     /label=chicken beta-actin promoter
     intron          658..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1682..1701
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1724..1733
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     repeat_region   1738..1772
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     CDS             complement(join(2032..2745,2746..2748))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VNKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2077..2098)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2426..2445
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2685..2706
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     repeat_region   2748..2810
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     3166..3185
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    3184..3239
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     3285..3304
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     rep_origin      complement(3451..4039)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3531..3550)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4210..5001,5002..5070))
                     /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     4833..4852
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5071..5175)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct
      421 cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga
      481 tgggggcggg gggggggggg gggcgcgccr gggggggggg gggggggggg ggggsggggs
      541 ggggsgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
      601 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
      661 gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc
      721 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
      781 ggctgtaatt agcgcttggt ttaatgacgg cttgtttctt ttctgtggct gcgtgaaagc
      841 cttgaggggc tccgggaggg ccctttgtgc ggggggagcg gctcgggggg tgcgtgcgtg
      901 tgtgtgtgcg tggggagcgc cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc
      961 gggcgcggcg cggggctttg tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc
     1081 gtgggggggt gagcaggggg tgtgggcgcg tcggtcgggc tgcaaccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggrggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aaggccacca tggttaaccc
     1741 tagaaagata gtctgcgtaa aattgacgca tgcatcaatt gtaactgcgg tttaaactcc
     1801 ccagcatgcc tgctattgtc ttcccaatcc tcccccttgc tgtcctgccc caccccaccc
     1861 cccagaatgg aatgacacct actcagacaa tgcgatgcaa tttcctcatt ttattaggaa
     1921 aggacagtga gagtggcacc ttccagggtc aaggaaggca cgggggaggg gcaaacaaca
     1981 gatggctggc aactagaagg cactaaagtc ggatccagag tcgcggccgc tttacttgta
     2041 cagctcgtcc atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg
     2101 atcgcgcttc tcgttggggt ctttgctcag ggcggactgg gtgctcaggt agtggttgtc
     2161 gggcagcagc acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg
     2221 cacgctgccg tcctcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg
     2281 cttgtcggcc atgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag
     2341 gatgttgccg tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc
     2401 gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt
     2461 gcgctcctgg acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg
     2521 gtcggggtag cggctgaagc actgcacgcc gtaggtcagg gtggtcacga gggtgggcca
     2581 gggcacgggc agcttgccgg tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc
     2641 atcgccctcg ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc
     2701 gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc ttattaaccc tagaaagata
     2761 atcatattgt gacgtacgtt aaagataatc atgcgtaaaa ttgacgcatg tccggagccg
     2821 tagatatcgg aaagaaagcg gccgctgcag gtcgagggat ctccataaga gaagagggac
     2881 agctatgact gggagtagtc aggagaggag gaaaaatctg gctagtaaaa catgtaagga
     2941 aaattttagg gatgttaaag aaaaaaataa cacaaaacaa aatataaaaa aaatctaacc
     3001 tcaagtcaag gcttttctat ggaataagga atggacagca gggggctgtt tcatatactg
     3061 atgacctctt tatagccaac ctttgttcat ggcagccagc atatgggcat atgttgccaa
     3121 actctaaacc aaatactcat tctgatgttt taaatgattt gccctcccat atgtccttcc
     3181 gagtgagaga cacaaaaaat tccaacacac tattgcaatg aaaataaatt tcctttatta
     3241 gccagaagtc agatgctcaa ggggcttcat gatgtcccca taatttttgg cagagggaaa
     3301 aagatctcag tggtatttgt gagccagggc attggccaca ccagccacca ccttctgata
     3361 ggcagcctgc acctgaggag tgaattagaa gctttgagca aaaggccagc aaaaggccag
     3421 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3481 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3541 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3601 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     3661 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     3721 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     3781 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     3841 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     3901 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     3961 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4021 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4081 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4141 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4201 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4261 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4321 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     4381 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4441 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4501 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     4561 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4621 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     4681 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     4741 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     4801 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     4861 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     4921 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     4981 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5041 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     5101 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5161 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcccaa tgatatcatt
     5221 taaatgtc
//
