LOCUS       sequence-289567-        6634 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of HsNxf-1-R233AR276A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6634)
  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 6634)
  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-R233AR276A_O
FEATURES             Location/Qualifiers
     source          1..6634
                     /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    3422..3543
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3459..3478)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3513..3532
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3550..4005)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3687..3708)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3899..3918
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4032..4136
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4138..4495
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4138..4329
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4161..4181)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4346..4481
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4530..5324
                     /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             4530..5324
                     /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(4584..4603)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5194..5213
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5512..5531)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5556..5603
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5932..6520
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6421..6440
                     /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 atatggcgga
     1441 cgaggggaag tcgtacagcg aacacgatga tgaacgcgtt aatttccctc aaagaaagaa
     1501 gaaaggccgg ggtcccttcc ggtggaaata tggtgaagga aaccgtaggt ctggaagagg
     1561 cggttctggt attcggtctt cccgccttga ggaagatgat ggagatgtgg caatgagtga
     1621 tgcccaggat ggtccccgag tacgatacaa cccctatacc acccgaccta accgtcgggg
     1681 tgatacttgg catgatcgag atcgcattca tgttactgtg cggagagaca gagctcctcc
     1741 agagagagga ggggctggca ccagccagga tgggacctca aagaactggt tcaagattac
     1801 aattccttat ggcagaaagt atgacaaggc atggctcctg agcatgattc agagcaagtg
     1861 cagtgtgccc ttcaccccta ttgagtttca ctatgagaat acacgggccc agttcttcgt
     1921 tgaagacgcc agtactgcct ctgcattgaa ggctgtcaac tataagattt tggatcggga
     1981 gaaccgaagg atatctatca tcatcaactc ttctgctcca ccccacacta tactgaatga
     2041 actgaagcca gaacaagtag aacagctaaa gctgatcatg agcaaacgat acgatggctc
     2101 ccaacaagcc cttgacctca aaggcctcgc ttcagaccca gatttggtgg cccagaacat
     2161 tgacgttgtc ctgaatcgca gaagctgtat ggcagctacc ctgaggatca ttgaagagaa
     2221 catccctgag ctattgtcct tgaacttgag caacaacgcg ctgtacaggc tggatgacat
     2281 gtctagcatt gttcagaagg cacccaacct gaagatccta aacctttctg gaaatgaatt
     2341 gaagtctgag cgggaattgg acaagataaa ggggctgaag ctagaagagc tctggctcga
     2401 tggaaactcc ctgtgtgaca ccttccgaga ccagtccacc tacatcagcg ccattcgcga
     2461 acgatttccc aagttactac gcctggatgg ccatgagcta cccccaccaa ttgcctttga
     2521 tgttgaagcc cccacgacgt taccgccctg caagggaagc tattttggaa cagaaaactt
     2581 gaagagtctg gtcttgcact tcctgcaaca gtactatgca atttacgact ctggagaccg
     2641 acaagggctc ctggatgcct accatgatgg ggcctgctgt tccctgagca ttcctttcat
     2701 tcctcagaac cctgcccgaa gcagcttagc cgagtatttc aaggatagca gaaatgtgaa
     2761 gaagcttaaa gaccctacct tgcggttccg gctgctgaag cacacgcgtc tcaacgttgt
     2821 tgccttcctc aatgagttgc ccaaaaccca gcacgacgtc aattccttcg tggtagacat
     2881 aagcgcccag acaagcacat tgctgtgttt ttctgtcaat ggagtcttca aggaagtgga
     2941 cggaaagtcc cgggattctt tgcgagcctt cacccggaca ttcattgctg ttcctgctag
     3001 caattcaggg ctatgtattg taaatgatga gctatttgtg cggaatgcca gttctgaaga
     3061 gatccaaaga gccttcgcta tgcctgcacc cacgccttcc tccagcccgg tgcccaccct
     3121 ctctccagag cagcaggaaa tgttgcaagc attctctacc cagtctggca tgaacctcga
     3181 gtggtcccag aagtgccttc aggacaacaa ctgggactac accagatctg cccaggcctt
     3241 cactcatctc aaggccaagg gcgagatccc agaagtggca ttcatgaagt gaggatccac
     3301 cggatctaga taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt
     3361 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     3421 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     3481 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     3541 ttaacgcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     3601 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     3661 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     3721 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     3781 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     3841 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     3901 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     3961 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc
     4021 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     4081 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg
     4141 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     4201 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     4261 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     4321 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     4381 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     4441 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaagatcg
     4501 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     4561 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     4621 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     4681 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
     4741 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     4801 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     4861 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     4921 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     4981 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     5041 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     5101 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     5161 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     5221 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     5281 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     5341 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     5401 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     5461 gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa ctgaaacacg
     5521 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     5581 gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct ggcactctgt
     5641 cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct tttccccacc
     5701 ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc
     5761 tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc atttttaatt
     5821 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     5881 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     5941 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     6001 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6061 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6121 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     6181 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6241 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6301 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6361 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6421 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6481 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6541 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     6601 tgattctgtg gataaccgta ttaccgccat gcat
//
