LOCUS       sequence-474969-        6628 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNxf-1-5xmut.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6628)
  AUTHORS   Teplova M, Wohlbold L, Khin NW, Izaurralde E, Patel DJ
  TITLE     Structure-function studies of nucleocytoplasmic transport of 
            retroviral genomic RNA by mRNA export factor TAP.
  JOURNAL   Nat Struct Mol Biol. 2011 Aug 7;18(9):990-8. doi: 10.1038/nsmb.2094.
  PUBMED    21822283
REFERENCE   2  (bases 1 to 6628)
  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: "10.1038/nsmb"; 
            journalName: "Nat Struct Mol Biol"; date: "2011-08-7- 7"; volume: 
            "18"; issue: "9"; pages: "990-8"
COMMENT     Sequence Label: pT7-EGFP-C1-HsNXF-1-5xmut_O
FEATURES             Location/Qualifiers
     source          1..6628
                     /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"
     primer_bind     613..632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        613..631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      659..668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(710..731)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(971..990)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1318..1339
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3416..3537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3453..3472)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3507..3526
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3544..3999)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3681..3702)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3893..3912
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4026..4130
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4132..4489
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4132..4323
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4155..4175)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4340..4475
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4524..5318
                     /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             4524..5318
                     /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(4578..4597)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5188..5207
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5506..5525)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5550..5597
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5926..6514
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6415..6434
                     /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 gcagagctct ctggctaact agagaaccca ctgcttactg
      601 gcttatcgaa attaatacga ctcactatag ggagacccaa gctctagcgc taccggtcgc
      661 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      721 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
      781 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
      841 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
      901 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
      961 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1021 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1081 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1141 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1201 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1261 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     1321 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1381 gtccggactc agatctcgag ctcaagcttc gaattctgca gtcgccatgg cggacgaggg
     1441 gaagtcgtac agcgaacacg atgatgaacg cgttaatttc cctcaaagaa agaagaaagg
     1501 ccggggtccc ttccggtgga aatatggtga aggaaaccgt aggtctggaa gaggcggttc
     1561 tggtattcgg tcttcccgcc ttgaggaaga tgatggagat gtggcaatga gtgatgccca
     1621 ggatggtccc cgagtacgat acaaccccta taccacccga cctaaccgtc ggggtgatac
     1681 ttggcatgat cgagatcgca ttcatgttac tgtgcggaga gacagagctc ctccagagag
     1741 aggaggggct ggcaccagcc aggatgggac ctcaaagaac tggttcaaga ttacaattcc
     1801 ttatggcgaa aagtatgaca aggcatggct cctgagcatg attcagagca agtgcggtgt
     1861 gcccttcacc cctattgagt ttcactatga gaatacagag gcccagttct tcgttgaaga
     1921 cgccagtact gcctctgcat tgaaggctgt caactataag attttggatc gggagaaccg
     1981 aaggatatct atcatcatca actcttctgc tccaccccac actatactga atgaactgaa
     2041 gccagaacaa gtagaacagc taaagctgat catgagcaaa cgatacgatg gctcccaaca
     2101 agcccttgac ctcaaaggcc tccgttcaga cccagatttg gtggcccaga acattgacgt
     2161 tgtcctgaat cgcagaagct gtatggcagc taccctgagg atcattgaag agaacatccc
     2221 tgagctattg tccttgaact tgagcaacaa caggctggcc gcgctggatg acatgtctag
     2281 cattgttcag aaggcaccca acctgaagat cctaaacctt tctggaaatg aattggcgtc
     2341 tgagcgggaa ttggacaaga taaaggggct gaagctagaa gagctctggc tcgatggaaa
     2401 ctccctgtgt gacaccttcc gagaccagtc cacctacatc agcgccattc gcgaacgatt
     2461 tcccaagtta ctacgcctgg atggccatga gctaccccca ccaattgcct ttgatgttga
     2521 agcccccacg acgttaccgc cctgcaaggg aagctatttt ggaacagaaa acttgaagag
     2581 tctggtcttg cacttcctgc aacagtacta tgcaatttac gactctggag accggcaagg
     2641 gctcctggat gcctaccatg atggggcctg ctgttccctg agcattcctt tcattcctca
     2701 gaaccctgcc cgaagcagct tagccgagta tttcaaggat agcagaaatg tgaagaagct
     2761 taaagaccct accttgcggt tccggctgct gaagcacacg cgtctcaacg ttgttgcctt
     2821 cctcaatgag ttgcccaaaa cccagcacga cgtcaattcc ttcgtggtag acataagcgc
     2881 ccagacaagc acattgctgt gtttttctgt caatggagtc ttcaaggaag tggacggaaa
     2941 gtcccgggat tctttgcgag ccttcacccg gacattcatt gctgttcctg ctagcaattc
     3001 agggctatgt attgtaaatg atgagctatt tgtgcggaat gccagttctg aagagatcca
     3061 aagagccttc gctatgcctg cacccacgcc ttcctccagc ccggtgccca ccctctctcc
     3121 agagcagcag gaaatgttgc aagcattctc tacccagtct ggcatgaacc tcgagtggtc
     3181 ccagaagtgc cttcaggaca acaactggga ctacaccaga tctgcccagg ccttcactca
     3241 tctcaaggcc aagggcgaga tcccagaagt ggcattcatg aagtgaggat ccaccggatc
     3301 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     3361 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     3421 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3481 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg
     3541 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     3601 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     3661 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     3721 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     3781 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     3841 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     3901 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     3961 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     4021 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     4081 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     4141 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     4201 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     4261 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     4321 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     4381 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     4441 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     4501 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     4561 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     4621 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     4681 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     4741 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     4801 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     4861 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     4921 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     4981 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     5041 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     5101 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     5161 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     5221 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     5281 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     5341 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     5401 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     5461 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     5521 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     5581 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     5641 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     5701 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     5761 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5821 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5881 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5941 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     6001 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6061 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6121 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6181 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6241 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6301 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6361 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6421 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     6481 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6541 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     6601 tgtggataac cgtattaccg ccatgcat
//
