LOCUS       sequence-76196-e        5831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a GFP-dependent transcription factor component in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5831)
  AUTHORS   Tang JC, Szikra T, Kozorovitskiy Y, Teixiera M, Sabatini BL, Roska 
            B, Cepko CL
  TITLE     A nanobody-based system using fluorescent proteins as scaffolds for 
            cell-specific gene manipulation.
  JOURNAL   Cell. 2013 Aug 15;154(4):928-39. doi: 10.1016/j.cell.2013.07.021.
  PUBMED    23953120
REFERENCE   2  (bases 1 to 5831)
  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.cell.2013.07"; journalName: "Cell"; date: "2013-08-15- 
            15"; volume: "154"; issue: "4"; pages: "928-39"
COMMENT     Sequence Label: pCAG-GBP1-10gly-lexADBD
FEATURES             Location/Qualifiers
     source          1..5831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1762..1771
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1777..2118
                     /codon_start=1
                     /product="GFP-binding fragment of a single-chain camelid 
                     antibody (Rothbauer et al., 2008)"
                     /label=GFP nanobody
                     /translation="VQLVESGGALVQPGGSLRLSCAASGFPVNRYSMRWYRQAPGKERE
                     WVAGMSSAGDRSSYEDSVKGRFTISRDDARNTVYLQMNSLKPEDTAVYYSNVNVGFEYW
                     GQGTQVTVSS"
     CDS             join(2161..2412,2413..2769)
                     /codon_start=1
                     /gene="lexA"
                     /product="E. coli DNA-binding protein that represses genes 
                     involved in the SOS response to DNA damage"
                     /label=LexA
                     /note="The N-terminal DNA-binding domain recognizes the 
                     16-bp palindromic sequence 5'-CTGTATATATATACAG-3'."
                     /translation="MKALTARQQEVFDLIRDHISQTGMPPTRAEIAQRLGFRSPNAAEE
                     HLKALARKGVIEIVSGASRGIRLLQEEEEGLPLVGRVAAGEPLLAQQHIEGHYQVDPSL
                     FKPNADFLLRVSGMSMKDIGIMDGDLLAVHKTQDVRNGQVVVARIDDEVTVKRLKKQGN
                     KVELLPENSEFKPIVVDLRQQSFTIEGLAVGVIRNGDWL"
     primer_bind     2704..2725
                     /label=LexA
                     /note="LexA DNA binding domain, forward primer"
     primer_bind     complement(2855..2874)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2920..2975
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2974..2993)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3334..3350)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3334..3350)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3347..3369)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3358..3374
                     /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(3382..3388,3389..3406,3407..3412))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3427..3448
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3507..3702
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3553..3688
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3708..3842
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3745..3764)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3799..3818
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3910..3927)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4081..4669)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4161..4180)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4840..5631,5632..5700))
                     /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     5463..5482
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5701..5805)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attctgcagt cgacggtacc
     1741 gcgggcccgg gatccaccgg tgccaccatg gccgacgtgc agctcgtgga atccggagga
     1801 gcactggtcc agcctggggg atcactgaga ctgtcttgtg ccgcttctgg ctttcctgtg
     1861 aatcggtact ctatgagatg gtacaggcag gcacctggaa aagagcgaga atgggtcgcc
     1921 ggaatgtcat ctgctggcga taggagttcc tacgaggata gcgtgaaagg acggttcacc
     1981 attagtaggg acgatgctag aaacaccgtg tacctccaga tgaactccct gaaacctgag
     2041 gacactgctg tgtactactc caacgtcaat gtgggctttg aatactgggg acagggcaca
     2101 caagtgactg ttagttctgc tagcggagga ggtggtggag gtggaggtgg aggtactagt
     2161 atgaaagcgt taacggccag gcaacaagag gtgtttgatc tcatccgtga tcacatcagc
     2221 cagacaggta tgccgccgac gcgtgcggaa atcgcgcagc gtttggggtt ccgttcccca
     2281 aacgcggctg aagaacatct gaaggcgctg gcacgcaaag gcgttattga aattgtttcc
     2341 ggcgcatcac gcgggattcg tctgttgcag gaagaggaag aagggttgcc gctggtaggt
     2401 cgtgtggctg ccggtgaacc acttctggcg caacagcata ttgaaggtca ttatcaggtc
     2461 gatccttcct tattcaagcc gaatgctgat ttcctgctgc gcgtcagcgg gatgtcgatg
     2521 aaagatatcg gcattatgga tggtgacttg ctggcagtgc ataaaactca ggatgtacgt
     2581 aacggtcagg tcgttgtcgc acgtattgat gacgaagtta ccgttaagcg cctgaaaaaa
     2641 cagggcaata aagtcgaact gttgccagaa aatagcgagt ttaaaccaat tgtcgttgac
     2701 cttcgtcagc agagcttcac cattgaaggt ctggcggttg gggttattcg caacggcgac
     2761 tggctgtaag cggccgcact cctcaggtgc aggctgccta tcagaaggtg gtggctggtg
     2821 tggccaatgc cctggctcac aaataccact gagatctttt tccctctgcc aaaaattatg
     2881 gggacatcat gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca
     2941 ttgcaatagt gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa
     3001 atcatttaaa acatcagaat gagtatttgg tttagagttt ggcaacatat gccatatgct
     3061 ggctgccatg aacaaaggtg gctataaaga ggtcatcagt atatgaaaca gccccctgct
     3121 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt
     3181 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga
     3241 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atgaagatcc
     3301 ctcgacctgc agcccaagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     3361 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     3421 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     3481 gggaaacctg tcgtgccagc ggatccgcat ctcaattagt cagcaaccat agtcccgccc
     3541 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3601 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     3661 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctaac ttgtttattg
     3721 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     3781 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     3841 tccgctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     3901 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     3961 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4021 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4081 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4141 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4201 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4261 cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4321 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4381 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4441 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4501 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
     4561 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4621 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4681 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4741 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     4801 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     4861 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     4921 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     4981 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5041 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     5101 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     5161 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5221 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5281 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     5341 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     5401 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     5461 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     5521 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     5581 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     5641 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     5701 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     5761 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     5821 agtgccacct g
//
