LOCUS       sequence-245170-        4832 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  a mammalian expression backbone vector for humanized 
            mNeonGreen-3xFlag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4832)
  AUTHORS   Tanida-Miyake E, Koike M, Uchiyama Y, Tanida I
  TITLE     Optimization of mNeonGreen for Homo sapiens increases its 
            fluorescent intensity in mammalian cells.
  JOURNAL   PLoS One. 2018 Jan 17;13(1):e0191108. doi: 
            10.1371/journal.pone.0191108. eCollection 2018.
  PUBMED    29342181
REFERENCE   2  (bases 1 to 4832)
  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: "PLoS One.";
            date: "2018-01-17"; volume: "13(1):e0191108. doi"; pages: " 
            10.1371/journal.pone.0191108. eCollection 2018"
COMMENT     Sequence Label: pmNeonGreenHO-3xFLAG-G
FEATURES             Location/Qualifiers
     source          1..4832
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1338..1361
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1374..1439
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    1440..1496
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1620..1741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1657..1676)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1711..1730
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1748..2203)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1885..1906)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2097..2116
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2230..2334
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2336..2693
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2336..2527
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2359..2379)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2544..2679
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2728..3522
                     /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             2728..3522
                     /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(2782..2801)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3392..3411
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3710..3729)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3754..3801
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4130..4718
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4619..4638
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcca
      601 ccatggtcag caaaggcgaa gaagacaaca tggcatcact gccagcgaca cacgaacttc
      661 atattttcgg ctccatcaat ggtgtggact tcgatatggt tggacagggt actgggaacc
      721 ctaacgatgg atatgaggag ttgaacttga agagtaccaa gggcgatctt cagttttcac
      781 cttggattct cgtacctcat ataggttatg gctttcatca atatctgccc tatcctgacg
      841 gtatgtcacc tttccaagca gctatggttg acgggagcgg ttatcaggtt caccggacaa
      901 tgcagtttga agatggagcc tctttgaccg taaattatcg gtatacttat gagggcagcc
      961 atattaaagg cgaggctcaa gttaagggga ctgggttccc cgcggatggt cccgtaatga
     1021 caaacagtct gacggctgca gactggtgcc gctctaagaa gacatatcca aatgacaaga
     1081 cgattattag cactttcaag tggtcataca ctacaggaaa tggtaaacgc tataggagca
     1141 ctgcccggac cacgtatact ttcgcgaagc cgatggcggc aaattatttg aaaaaccaac
     1201 cgatgtatgt gttcagaaag acagaactca agcattctaa aactgaactg aacttcaaag
     1261 agtggcagaa agcctttact gatgtaatgg gcatggacga actctataag tccggaggtg
     1321 gatcgggagg tggatcgtgg agccacccgc agttcgaaaa aactagtcgg gctgactaca
     1381 aagaccatga cggtgattat aaagatcatg acatcgacta caaggatgac gatgacaaga
     1441 gatctcgagc tcaagcttcg aattctgcag tcgacggtac cgcgggcccg ggatccaccg
     1501 gatctagata actgatcata atcagccata ccacatttgt agaggtttta cttgctttaa
     1561 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     1621 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     1681 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     1741 aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     1801 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     1861 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     1921 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     1981 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     2041 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     2101 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     2161 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg
     2221 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     2281 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtcctgag
     2341 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     2401 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     2461 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2521 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     2581 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     2641 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaagatcgat
     2701 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     2761 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     2821 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     2881 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     2941 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     3001 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     3061 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     3121 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     3181 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     3241 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     3301 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     3361 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     3421 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     3481 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg
     3541 aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct
     3601 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     3661 gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact gaaacacgga
     3721 aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag aataaaacgc
     3781 acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     3841 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     3901 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     3961 ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat ttttaattta
     4021 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     4081 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4141 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     4201 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     4261 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     4321 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     4381 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     4441 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     4501 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     4561 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     4621 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     4681 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     4741 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     4801 attctgtgga taaccgtatt accgccatgc at
//
