LOCUS       sequence-198192-        5506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of human VAP-B (mutant K87D/M89D, unable to bind FFAT 
            motifs) fused to GFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5506)
  AUTHORS   Alpy F, Rousseau A, Schwab Y, Legueux F, Stoll I, Wendling C, 
            Spiegelhalter C, Kessler P, Mathelin C, Rio MC, Levine TP, Tomasetto
            C
  TITLE     STARD3 or STARD3NL and VAP form a novel molecular tether between 
            late endosomes and the ER.
  JOURNAL   J Cell Sci. 2013 Dec 1;126(Pt 23):5500-12. doi: 10.1242/jcs.139295. 
            Epub 2013 Oct 8.
  PUBMED    24105263
REFERENCE   2  (bases 1 to 5506)
  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: "J Cell 
            Sci."; date: "2013-12-1"; volume: "126(Pt 23):5500-12. doi"; pages: 
            " 10.1242/jcs.139295. Epub 2013 Oct 8"
COMMENT     Sequence Label: pEGFPC1-hVAP-B KD/MD
FEATURES             Location/Qualifiers
     source          1..5506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        57..360
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        361..564
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     515..535
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      603..612
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(609..611,612..614,615..1325)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(609..611,612..614,615..1325)
                     /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(654..675)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(915..934)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1262..1283
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1365..1391
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    2290..2411
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2327..2346)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2381..2400
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2418..2873)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2555..2576)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2767..2786
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2900..3004
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3006..3363
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3006..3197
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3029..3049)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3214..3349
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3398..4192
                     /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             3398..4192
                     /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(3452..3471)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4062..4081
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4380..4399)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4424..4471
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4800..5388
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5289..5308
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
       61 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      121 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      181 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      241 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      301 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      361 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      421 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      481 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      541 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
      601 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      661 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
      721 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      781 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
      841 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
      901 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
      961 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1021 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     1081 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     1141 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1201 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1261 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1321 acaagtccgg actcagatct cgagctcaag cttcgaattg tgcctaccca tatgacgtcc
     1381 cggactacgc cggatccatg gcgaaggtgg agcaggtcct gagcctcgag ccgcagcacg
     1441 agctcaaatt ccgaggtccc ttcaccgatg ttgtcaccac caacctaaag cttggcaacc
     1501 cgacagaccg aaatgtgtgt tttaaggtga agactacagc accacgtagg tactgtgtga
     1561 ggcccaacag cggaatcatc gatgcagggg cctcaattaa tgtatctgtg atgttacagc
     1621 ctttcgatta tgatcccaat gagaaaagta aacacgactt tgacgtccag tctatgtttg
     1681 ctccaactga cacttcagat atggaagcag tatggaagga ggcaaaaccg gaagacctta
     1741 tggattcaaa acttagatgt gtgtttgaat tgccagcaga gaatgataaa ccacatgatg
     1801 tagaaataaa taaaattata tccacaactg catcaaagac agaaacacca atagtgtcta
     1861 agtctctgag ttcttctttg gatgacaccg aagttaagaa ggttatggaa gaatgtaaga
     1921 ggctgcaagg tgaagttcag aggctacggg aggagaacaa gcagttcaag gaagaagatg
     1981 gactgcggat gaggaagaca gtgcagagca acagccccat ttcagcatta gccccaactg
     2041 ggaaggaaga aggccttagc acccggctct tggctctggt ggttttgttc tttatcgttg
     2101 gtgtaattat tgggaagatt gccttgtagc aattctgcag tcgacggtac cgcgggcccg
     2161 ggatccaccg gatctagata actgatcata atcagccata ccacatttgt agaggtttta
     2221 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     2281 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     2341 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     2401 aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     2461 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     2521 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     2581 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     2641 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     2701 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     2761 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     2821 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg
     2881 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2941 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3001 gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3061 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3121 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3181 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3241 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3301 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3361 aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     3421 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     3481 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     3541 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc
     3601 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     3661 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     3721 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     3781 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     3841 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     3901 cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca
     3961 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     4021 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     4081 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     4141 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     4201 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     4261 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     4321 atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact
     4381 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     4441 aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     4501 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     4561 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     4621 gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat
     4681 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4741 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4801 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4861 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4921 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4981 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     5041 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     5101 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     5161 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     5221 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     5281 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     5341 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     5401 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     5461 ttatcccctg attctgtgga taaccgtatt accgccatgc attagt
//
