LOCUS       sequence-313671-        4687 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Backbone for making protein-responsive mRNA switches.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4687)
  AUTHORS   Kawasaki S, Fujita Y, Nagaike T, Tomita K, Saito H
  TITLE     Synthetic mRNA devices that detect endogenous proteins and 
            distinguish mammalian cells.
  JOURNAL   Nucleic Acids Res. 2017 May 19. doi: 10.1093/nar/gkx298.
  PUBMED    28525643
REFERENCE   2  (bases 1 to 4687)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2017 May 19. doi: 10.1093/nar/gkx298."
COMMENT     Sequence Label: pAptamerCassette-EGFP
FEATURES             Location/Qualifiers
     source          1..4687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      567..576
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(573..575,576..578,579..1292)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(573..575,576..578,579..1289)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(618..639)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(879..898)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1226..1247
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1415..1536
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1452..1471)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1506..1525
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1543..1998)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1680..1701)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1892..1911
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2025..2129
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2131..2488
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2131..2322
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2154..2174)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2339..2474
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2523..3317
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2523..3317
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2577..2596)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3187..3206
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3505..3524)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3549..3596
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3925..4513
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4414..4433
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcggat
      541 cctaatacga ctcactataa ccggtcgcca ccatggtgag caagggcgag gagctgttca
      601 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
      661 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
      721 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
      781 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
      841 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
      901 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
      961 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1021 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1081 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1141 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     1201 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     1261 tcactctcgg catggacgag ctgtacaagt aaagcggccg cgactctaga tcataatcag
     1321 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     1381 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     1441 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     1501 tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata
     1561 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1621 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1681 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1741 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1801 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     1861 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     1921 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     1981 cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     2041 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2101 tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt
     2161 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     2221 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     2281 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     2341 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2401 tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg
     2461 cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc
     2521 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     2581 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     2641 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     2701 tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     2761 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     2821 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     2881 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     2941 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3001 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg
     3061 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     3121 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     3181 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     3241 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     3301 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     3361 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3421 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     3481 cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg
     3541 cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa
     3601 acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg
     3661 ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc
     3721 agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat
     3781 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     3841 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     3901 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     3961 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     4021 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     4081 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     4141 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     4201 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     4261 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     4321 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     4381 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     4441 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     4501 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     4561 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     4621 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
     4681 agttccg
//
