LOCUS       sequence-338279-        4819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing EGFP mRNA fused with 2 tandem repeats of a 50-base 
            sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4819)
  AUTHORS   Chen M, Ma Z, Wu X, Mao S, Yang Y, Tan J, Krueger CJ, Chen AK
  TITLE     A molecular beacon-based approach for live-cell imaging of RNA 
            transcripts with minimal target engineering at the single-molecule 
            level.
  JOURNAL   Sci Rep. 2017 May 8;7(1):1550. doi: 10.1038/s41598-017-01740-1.
  PUBMED    28484218
REFERENCE   2  (bases 1 to 4819)
  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"; journalName: "Sci Rep."; 
            date: "2017-05-8"; volume: "7(1):1550. doi"; pages: " 
            10.1038/s41598-017-01740-1"
COMMENT     Sequence Label: pEGFP-N1-2x
FEATURES             Location/Qualifiers
     source          1..4819
                     /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      699..708
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(705..707,708..710,711..1424)
                     /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(750..771)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1011..1030)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1358..1379
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1547..1668
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1584..1603)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1638..1657
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1675..2130)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1812..1833)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2024..2043
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2157..2261
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2263..2620
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2263..2454
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2286..2306)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2471..2606
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2655..3449
                     /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             2655..3449
                     /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(2709..2728)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3319..3338
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3637..3656)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3681..3728
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4057..4645
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4546..4565
                     /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 gctagcgcta
      541 ccggactcag atctcgagct caagcttcga attcgtcgac aggagttgtg tttgtggacg
      601 aagagcacca gccagctgat cgacctcgac aggagttgtg tttgtggacg aagagcacca
      661 gccagctgat cgacctcgag ccgagggatc caccggtcgc caccatggtg agcaagggcg
      721 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
      781 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
      841 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
      901 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
      961 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1021 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1081 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     1141 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     1201 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     1261 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     1321 ccaagctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     1381 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcgactcta
     1441 gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca
     1501 cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
     1561 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     1621 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg
     1681 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     1741 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     1801 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     1861 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     1921 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     1981 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     2041 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     2101 ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg
     2161 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2221 aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca
     2281 gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     2341 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     2401 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     2461 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     2521 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     2581 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat
     2641 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     2701 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     2761 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     2821 ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     2881 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     2941 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3001 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     3061 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     3121 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     3181 cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     3241 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     3301 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     3361 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     3421 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     3481 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     3541 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     3601 gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc
     3661 ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc
     3721 gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag
     3781 accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc
     3841 gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg
     3901 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     3961 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4021 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4081 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4141 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     4201 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4261 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4321 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4381 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     4441 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     4501 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4561 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4621 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4681 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     4741 ccgtattacc gccatgcatt agttattaat agtaatcaat tacggggtca ttagttcata
     4801 gcccatatat ggagttccg
//
