LOCUS       sequence-384656-        5141 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mEGFP mRNA with 2xbiRhoBAST in 3'UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5141)
  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 5141)
  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-2xbiRhoBAST
FEATURES             Location/Qualifiers
     source          1..5141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        129..320
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     277..297
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(322..426)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      453..908
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(540..559)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     750..771
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    915..1036
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(926..945)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     980..999
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    1160..1212
                     /label=MCS
                     /note="multiple cloning site"
     CDS             complement(join(1639..2349,2350..2352,2353..2355))
                     /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(1681..1702)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2030..2049
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2289..2310
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2352..2361
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2397..2600)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2426..2446)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2601..2904
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3079..3667)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3159..3178)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3996..4043
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4068..4087
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4275..5069)
                     /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             complement(4275..5069)
                     /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(4386..4405)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4996..5015
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(5104..5141,1..320))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(5118..5141,1..112)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 acttctggaa tagctcagag gccgaggcgg cctcggcctc tgcataaata aaaaaaatta
       61 gtcagccatg gggcggagaa tgggcggaac tgggcggagt taggggcggg atgggcggag
      121 ttaggggcgg gactatggtt gctgactaat tgagatgcat gctttgcata cttctgcctg
      181 ctggggagcc tggggacttt ccacacctgg ttgctgacta attgagatgc atgctttgca
      241 tacttctgcc tgctggggag cctggggact ttccacaccc taactgacac acattccaca
      301 gctggttctt tccgcctcag gactcttcct ttttcaatat tattgaagca tttatcaggg
      361 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      421 tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc tgtagcggcg cattaagcgc
      481 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
      541 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
      601 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
      661 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
      721 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
      781 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
      841 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
      901 tacaatttac gcgttaagat acattgatga gtttggacaa accacaacta gaatgcagtg
      961 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     1021 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     1081 ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtatgg ctgattatga
     1141 tcagttatct agatccggtg gatcccgggc ccgcggtacc gtcgactgca gaattcgaag
     1201 cttgagctcg agatcgatgc ggccgccata tggatatcgg taaccgacgt caagcttgca
     1261 tgcctgcagg tcctagtaaa aagcggaccg aagtccgctc tagcggagcc cggataaggc
     1321 ctgcgccagt taaccttgtg gcgctccttc accgctttcg cggaggcctt tgcggagcga
     1381 actccgctgg aacctgaggc ggttaacctt gcgcctctcc ttcaccgctt tcgcggaggt
     1441 tccttccggg ctcctctagc ggtatgtggg taaggcctgc gccagttaac cttgtggcgc
     1501 tccttcaccg ctttcgcgga ggcctttgga ggacgaatcc tcctggaacc tgaggcggtt
     1561 aaccttgcgc ctctccttca ccgctttcgc ggaggttcct cccacatacc tctagacccg
     1621 tcgagatctg agtccggact tgtacagctc gtccatgccg agagtgatcc cggcggcggt
     1681 cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagcttgga
     1741 ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt
     1801 ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa
     1861 gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta
     1921 gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag
     1981 ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt
     2041 gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt
     2101 gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt
     2161 cagggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt
     2221 cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg
     2281 gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc
     2341 gcccttgctc accatggtgg cggaattcgc tagcggatct gacggttcac taaaccagct
     2401 ctgcttatat agacctccca ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag
     2461 ttgttacgac attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg
     2521 tcaatggggt ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattgatg
     2581 tactgccaaa accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa
     2641 gtaggaaagt cccataaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca
     2701 ttgacgtcaa tagggggcgt acttggcata tgatacactt gatgtactgc caagtgggca
     2761 gtttaccgta aatactccac ccattgacgt caatggaaag tccctattgg cgttactatg
     2821 ggaacatacg tcattattga cgtcaatggg cgggggtcgt tgggcggtca gccaggcggg
     2881 ccatttaccg taagttatgt aacgcggaac tccatatatg ggctatgaac taatgacccc
     2941 gtaattgatt actattaata actaatgcat ggcggtaata cggttatcca cagaatcagg
     3001 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     3061 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     3121 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     3181 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     3241 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     3301 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3361 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3421 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3481 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     3541 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3601 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3661 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3721 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3781 ttaaaaatga agttttaaat caatctaaag tatatatgag taacctgagg ctatggcagg
     3841 gcctgccgcc ccgacgttgg ctgcgagccc tgggccttca cccgaacttg gggggtgggg
     3901 tggggaaaag gaagaaacgc gggcgtattg gccccaatgg ggtctcggtg gggtatcgac
     3961 agagtgccag ccctgggacc gaaccccgcg tttatgaaca aacgacccaa caccgtgcgt
     4021 tttattctgt ctttttattg ccgtcatagc gcgggttcct tccggtattg tctccttccg
     4081 tgtttcagtt agcctccccc tagggtgggc gaagaactcc agcatgagat ccccgcgctg
     4141 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     4201 ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgaa
     4261 ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa
     4321 tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct
     4381 tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg
     4441 ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca
     4501 tcgccatggg tcacgacgag atcctcgccg tcgggcatgc tcgccttgag cctggcgaac
     4561 agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg
     4621 gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag
     4681 gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg
     4741 gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag
     4801 tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc cgtcgtggcc
     4861 agccacgata gccgcgctgc ctcgtcttgc agttcattca gggcaccgga caggtcggtc
     4921 ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc atcagagcag
     4981 ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc ggccggagaa
     5041 cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt ctcttgatcg
     5101 atctttgcaa aagcctaggc ctccaaaaaa gcctcctcac t
//
