LOCUS       sequence-190530-        5423 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EGFP-mCherry control reporter for methionine misincorporation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5423)
  AUTHORS   Gomes AC, Kordala AJ, Strack R, Wang X, Geslain R, Delaney K, Clark 
            WC, Keenan R, Pan T
  TITLE     A dual fluorescent reporter for the investigation of methionine 
            mistranslation in live cells.
  JOURNAL   RNA. 2016 Mar;22(3):467-76. doi: 10.1261/rna.054163.115. Epub 2016 
            Jan 4.
  PUBMED    26729921
REFERENCE   2  (bases 1 to 5423)
  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: 
            "10.1261/rna.054163.115"; journalName: "RNA"; date: "2016-03"; 
            volume: "22"; issue: "3"; pages: "467-76"
COMMENT     Sequence Label: WT-Pan
FEATURES             Location/Qualifiers
     source          1..5423
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        120..423
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        424..627
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     578..598
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      694..703
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(700..702,703..705,706..1416)
                     /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(745..766)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1006..1025)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1353..1374
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1432..1441
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1438..2148
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1583..1601)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1851..1870
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2270..2391
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2307..2326)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2361..2380
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2398..2853)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2535..2556)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2747..2766
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2880..2984
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2986..3343
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2986..3177
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3009..3029)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3194..3329
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3378..4172
                     /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             3378..4172
                     /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(3432..3451)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4042..4061
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4360..4379)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4404..4451
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4780..5368
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5269..5288
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt
       61 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
      121 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
      181 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      241 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      301 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
      361 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
      421 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
      481 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
      541 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
      601 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac
      661 cggactcaga tctcgagctc aagcttcgaa ttcgccacca tggtgagcaa gggcgaggag
      721 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
      781 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
      841 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
      901 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
      961 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1021 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1081 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1141 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1201 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1261 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1321 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1381 gccgggatca ctctcggcat ggacgagctg tacaagtcgg atccactagt cgccaccatg
     1441 gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg
     1501 cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
     1561 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc
     1621 gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc
     1681 gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
     1741 atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc
     1801 gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg
     1861 cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc
     1921 ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag
     1981 gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac
     2041 atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc
     2101 gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtaagc ggccgcgact
     2161 ctagatcata atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc
     2221 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat
     2281 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     2341 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa
     2401 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     2461 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     2521 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     2581 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     2641 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     2701 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     2761 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     2821 ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc
     2881 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     2941 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa
     3001 ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     3061 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     3121 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     3181 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     3241 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     3301 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag
     3361 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     3421 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     3481 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     3541 gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     3601 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3661 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     3721 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     3781 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     3841 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     3901 aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     3961 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     4021 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     4081 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     4141 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     4201 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4261 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4321 catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat
     4381 accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg
     4441 gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc
     4501 gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag
     4561 ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc
     4621 aggttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     4681 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcct
     4741 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     4801 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     4861 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     4921 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     4981 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5041 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5101 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5161 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5221 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5281 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5341 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5401 ggccttttgc tggccttttg ctc
//
