LOCUS       sequence-384642-        4985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mEGFP mRNA with biRhoBAST in 3'UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4985)
  AUTHORS   Buhler B, Schokolowski J, Benderoth A, Englert D, Grun F, Jaschke A,
            Sunbul M
  TITLE     Avidity-based bright and photostable light-up aptamers for 
            single-molecule mRNA imaging.
  JOURNAL   Nat Chem Biol. 2023 Apr;19(4):478-487. doi: 
            10.1038/s41589-022-01228-8. Epub 2023 Jan 19.
  PUBMED    36658339
REFERENCE   2  (bases 1 to 4985)
  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/s41589-022-01228-8"; journalName: "Nat Chem Biol"; date: 
            "2023-04"; volume: "19"; issue: "4"; pages: "478-487"
COMMENT     Sequence Label: mEGFP-biRhoBAST
FEATURES             Location/Qualifiers
     source          1..4985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        133..436
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        437..640
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     591..611
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      676..685
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(682..684,685..687,688..1398)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(727..748)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(988..1007)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1335..1356
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1669..1721
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1845..1966
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1882..1901)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1936..1955
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1973..2428)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2110..2131)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2322..2341
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2455..2559
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2561..2918
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2561..2752
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2584..2604)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2769..2904
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2953..3747
                     /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             2953..3747
                     /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(3007..3026)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3617..3636
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3935..3954)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3979..4026
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4355..4943
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4844..4863
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
       61 accgccatgc attagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      121 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      181 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      241 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      301 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      361 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      421 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      481 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      541 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      601 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
      661 ccgctagcga attccgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
      721 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
      781 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
      841 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
      901 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
      961 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1021 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1081 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     1141 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     1201 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1261 gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac
     1321 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1381 atggacgagc tgtacaagtc cggactcaga tctcgacggg tctagaggta tgtgggagga
     1441 acctccgcga aagcggtgaa ggagaggcgc aaggttaacc gcctcaggtt ccaggaggat
     1501 tcgtcctcca aaggcctccg cgaaagcggt gaaggagcgc cacaaggtta actggcgcag
     1561 gccttaccca cataccgcta gagcggactt cggtccgctt tttactagga cctgcaggca
     1621 tgcaagcttg acgtcggtta ccgatatcca tatggcggcc gcatcgatct cgagctcaag
     1681 cttcgaattc tgcagtcgac ggtaccgcgg gcccgggatc caccggatct agataactga
     1741 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     1801 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     1861 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     1921 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta
     1981 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     2041 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     2101 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     2161 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     2221 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     2281 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     2341 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     2401 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga
     2461 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     2521 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc
     2581 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2641 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2701 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     2761 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2821 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt
     2881 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga
     2941 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3001 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3061 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3121 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3181 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3241 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     3301 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     3361 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     3421 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     3481 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     3541 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     3601 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     3661 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     3721 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     3781 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     3841 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     3901 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg
     3961 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt
     4021 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac
     4081 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg
     4141 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt
     4201 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     4261 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     4321 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     4381 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     4441 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     4501 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     4561 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     4621 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     4681 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     4741 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     4801 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     4861 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     4921 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     4981 tttgc
//
