LOCUS       sequence-321663-        6306 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 
            5 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 6306)
  TITLE     Simon lab plasmids
REFERENCE   2  (bases 1 to 6306)
  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(0)
FEATURES             Location/Qualifiers
     source          1..6306
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        69..372
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        373..576
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     527..547
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    599..679
                     /label=MCS
                     /note="multiple cloning site"
     primer_bind     complement(2215..2236)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2476..2495)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2635..2886
                     /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     2823..2844
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3102..3223
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3139..3158)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3193..3212
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3230..3685)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3367..3388)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3579..3598
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3712..3816
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3818..4175
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3818..4009
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3841..3861)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4026..4161
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4210..5004
                     /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             4210..5004
                     /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(4264..4283)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4874..4893
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5192..5211)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5236..5283
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5612..6200
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6101..6120
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccatgcatta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
       61 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      121 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      181 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      241 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      301 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      361 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      421 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      481 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      541 aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc
      601 tagcgctacc ggactcagat ctcgagctca agcttcgaat tctgcagtcg acggtaccgc
      661 gggcccggga tccaccggtc ccgcgacatg gcattcaatt ttggggctcc ttcgggcact
      721 tcgggcacct ctacagccac cgcggcaccg gcgggcgggt ttggcggatt tggaacaaca
      781 actacaactg caggctctgc attcagcttt tctgccccaa caaacacagg cagtacaggc
      841 cttctcggcg gcactcagaa caaaggtttt ggctttggca ctggttttgg cacatcgacg
      901 ggtactggca ctggtttagg cactggcttg ggaaccggac ttggattcgg aggatttaac
      961 acccagcagc agcagcagca gcagcagact tctttaggcg gtctcttcag tcagcctgca
     1021 caggcccctg cgcagtccaa ccaactcatc aacactgcca gcgctctttc tgctccaacg
     1081 ctattggggg atgagagaga tgccatcttg gcaaagtgga accagttgca ggccttctgg
     1141 ggaacaggaa aaggatattt caataataac attcctccag tggagtttac acaggaaaac
     1201 cccttttgcc gatttaaggc tgtcggttac agttgtatgc ccaataacaa agatgaagat
     1261 ggactagtgg ttctaatttt caacaaaaaa gaaacagata ttagaagcca acaacagcag
     1321 ttggtagaat cattgcataa agttctggga ggaaaccaga ccctcactgt caatgtggag
     1381 ggcattaaga cactgcccga cgatcagaca gaagttgtca tttacattgt tgagcgctct
     1441 ccaaatggca cctcgagaag agttccagcg acaacgctgt atgcccactt tgagcaagcc
     1501 aacattaaaa cacagcttca gcagcttggc gtgacccttt ctatgaccag aacagagctg
     1561 tctcctgctc agatcaagca gctcctacag aaccctcctg ctggtgttga tcccattatt
     1621 tgggaacagg ccaaggtgga taacccggat tctgaaaaat taattcctgt tccaatggtg
     1681 ggtttcaaag aattgcttcg aagactgaag gttcaagacc agatgactaa gcagcaccag
     1741 accagattag atatcatatc tgaagatatt agtgaactgc aaaagaatca aactacaact
     1801 atggccaaaa tagcacaata caagaggaaa ctgatggatc tctcacacag aaccttacag
     1861 gtcctaatca agcaggaaat ccagaggaag agtggctatg ccatccaggc tgaggaagaa
     1921 cagttgcggg ttcagctaga tacgattcaa ggagagctga atgcacccac tcagttcaaa
     1981 ggtcgactga atgaactgat gtcccagatc aggatgcaga atcattttgg agctgtgaaa
     2041 tctgaagaaa agtattacat agatgcagac ctattacgag agatcaagca gcatttgaaa
     2101 cagcaacagg aaggccttag ccacctgatt agcatcataa aagacgacct ggaagatatc
     2161 aagctggtag aagctgctgc aaaggaggag ctgttcaccg gggtggtgcc catcctggtc
     2221 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     2281 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     2341 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
     2401 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     2461 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     2521 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     2581 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     2641 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     2701 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     2761 gacaaccact acctgagcac ccagtccaag ctgagcaaag accccaacga gaagcgcgat
     2821 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     2881 tacaaggagc atggactgaa tgagaccatc cacagcagag gcggggtctt tagctgacca
     2941 ttgcgctgtc cagaggttca taaagctgtg tacaagtaaa gcggccgcga ctctagatca
     3001 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     3061 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     3121 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3181 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     3241 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     3301 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     3361 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     3421 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     3481 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     3541 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     3601 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     3661 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     3721 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     3781 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     3841 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3901 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3961 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     4021 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     4081 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     4141 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     4201 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     4261 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4321 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4381 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4441 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4501 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4561 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4621 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4681 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     4741 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4801 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4861 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4921 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4981 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     5041 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     5101 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     5161 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     5221 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     5281 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     5341 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     5401 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     5461 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5521 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5581 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5641 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5701 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5761 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5821 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5881 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5941 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6001 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6061 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     6121 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     6181 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     6241 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     6301 ttaccg
//
