LOCUS       sequence-321664-        6309 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Nup54 with a mEGFP (missing the first 5 
            amino acids) at amino acid position 494 with a rigid alpha helix of 
            6 amino acids at the carboxyl end of the mEGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6309)
  TITLE     Simon lab plasmids
REFERENCE   2  (bases 1 to 6309)
  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"
COMMENT     Sequence Label: Nup54-mEGFP494(1)
FEATURES             Location/Qualifiers
     source          1..6309
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        70..373
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        374..577
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     528..548
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    600..680
                     /label=MCS
                     /note="multiple cloning site"
     primer_bind     complement(2219..2240)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2480..2499)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2639..2890
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNH
                     YLSTQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     2827..2848
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3106..3227
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3143..3162)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3197..3216
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3234..3689)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3371..3392)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3583..3602
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3716..3820
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3822..4179
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3822..4013
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3845..3865)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4030..4165
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4214..5008
                     /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             4214..5008
                     /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(4268..4287)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4878..4897
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5196..5215)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5240..5287
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5616..6204
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6105..6124
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gccatgcatt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
       61 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      121 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      181 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      241 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      301 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      361 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      421 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      481 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      541 taggcgtgta cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg
      601 ctagcgctac cggactcaga tctcgagctc aagcttcgaa ttctgcagtc gacggtaccg
      661 cgggcccggg atccaccggt cccgcgacat ggcattcaat tttggggctc cttcgggcac
      721 ttcgggcacc tctacagcca ccgcggcacc ggcgggcggg tttggcggat ttggaacaac
      781 aactacaact gcaggctctg cattcagctt ttctgcccca acaaacacag gcagtacagg
      841 ccttctcggc ggcactcaga acaaaggttt tggctttggc actggttttg gcacatcgac
      901 gggtactggc actggtttag gcactggctt gggaaccgga cttggattcg gaggatttaa
      961 cacccagcag cagcagcagc agcagcagac ttctttaggc ggtctcttca gtcagcctgc
     1021 acaggcccct gcgcagtcca accaactcat caacactgcc agcgctcttt ctgctccaac
     1081 gctattgggg gatgagagag atgccatctt ggcaaagtgg aaccagttgc aggccttctg
     1141 gggaacagga aaaggatatt tcaataataa cattcctcca gtggagttta cacaggaaaa
     1201 ccccttttgc cgatttaagg ctgtcggtta cagttgtatg cccaataaca aagatgaaga
     1261 tggactagtg gttctaattt tcaacaaaaa agaaacagat attagaagcc aacaacagca
     1321 gttggtagaa tcattgcata aagttctggg aggaaaccag accctcactg tcaatgtgga
     1381 gggcattaag acactgcccg acgatcagac agaagttgtc atttacattg ttgagcgctc
     1441 tccaaatggc acctcgagaa gagttccagc gacaacgctg tatgcccact ttgagcaagc
     1501 caacattaaa acacagcttc agcagcttgg cgtgaccctt tctatgacca gaacagagct
     1561 gtctcctgct cagatcaagc agctcctaca gaaccctcct gctggtgttg atcccattat
     1621 ttgggaacag gccaaggtgg ataacccgga ttctgaaaaa ttaattcctg ttccaatggt
     1681 gggtttcaaa gaattgcttc gaagactgaa ggttcaagac cagatgacta agcagcacca
     1741 gaccagatta gatatcatat ctgaagatat tagtgaactg caaaagaatc aaactacaac
     1801 tatggccaaa atagcacaat acaagaggaa actgatggat ctctcacaca gaaccttaca
     1861 ggtcctaatc aagcaggaaa tccagaggaa gagtggctat gccatccagg ctgaggaaga
     1921 acagttgcgg gttcagctag atacgattca aggagagctg aatgcaccca ctcagttcaa
     1981 aggtcgactg aatgaactga tgtcccagat caggatgcag aatcattttg gagctgtgaa
     2041 atctgaagaa aagtattaca tagatgcaga cctattacga gagatcaagc agcatttgaa
     2101 acagcaacag gaaggcctta gccacctgat tagcatcata aaagacgacc tggaagatat
     2161 caagctggta gcagaagctg ctgcaaagga ggagctgttc accggggtgg tgcccatcct
     2221 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     2281 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     2341 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     2401 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     2461 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     2521 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     2581 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     2641 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     2701 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     2761 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     2821 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     2881 gctgtacaag gagcatggac tgaatgagac catccacagc agaggcgggg tctttagctg
     2941 accattgcgc tgtccagagg ttcataaagc tgtgtacaag taaagcggcc gcgactctag
     3001 atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac
     3061 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
     3121 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     3181 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg cgtaaattgt
     3241 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     3301 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     3361 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     3421 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     3481 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     3541 tagagcttga cggggaaagt cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     3601 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     3661 cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg
     3721 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3781 accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag
     3841 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3901 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     3961 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     4021 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     4081 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     4141 tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg
     4201 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     4261 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     4321 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     4381 cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     4441 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     4501 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     4561 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     4621 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     4681 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     4741 gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     4801 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     4861 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     4921 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     4981 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     5041 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     5101 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     5161 ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg
     5221 gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg
     5281 tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga
     5341 ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg
     5401 ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt
     5461 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5521 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     5581 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5641 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5701 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5761 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5821 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5881 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     5941 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6001 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6061 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6121 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6181 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6241 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6301 cgtattacc
//
