LOCUS       sequence-198185-        5485 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of human VAP-A (mutant K94D/M96D, 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 5485)
  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 5485)
  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-A KD/MD
FEATURES             Location/Qualifiers
     source          1..5485
                     /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"
     polyA_signal    2269..2390
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2306..2325)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2360..2379
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2397..2852)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2534..2555)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2746..2765
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2879..2983
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2985..3342
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2985..3176
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3008..3028)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3193..3328
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3377..4171
                     /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             3377..4171
                     /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(3431..3450)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4041..4060
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4359..4378)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4403..4450
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4779..5367
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5268..5287
                     /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 tgcgtccgcc tcaggggcca
     1381 tggcgaagca cgagcagatc ctggtcctcg atccgcccac agacctcaaa ttcaaaggcc
     1441 ccttcacaga tgtagtcact acaaatctta aattgcgaaa tccatcggat agaaaagtgt
     1501 gtttcaaagt gaagactaca gcacctcgcc ggtactgtgt gaggcccaac agtggaatta
     1561 ttgacccagg gtcaactgtg actgtttcag taatgctaca gccctttgac tatgatccga
     1621 atgaaaagag taaacacgac tttgacgtcc agacaatttt tgctccacca aacacttcag
     1681 atatggaagc tgtgtggaaa gaggcaaaac ctgatgaatt aatggattcc aaattgagat
     1741 gcgtatttga aatgcccaat gaaaatgata aattgaatga tatggaacct agcaaagctg
     1801 ttccactgaa tgcatctaag caagatggac ctatgccaaa accacacagt gtttcactta
     1861 atgataccga aacaaggaaa ctaatggaag agtgtaaaag acttcaggga gaaatgatga
     1921 agctatcaga agaaaatcgg cacctgagag atgaaggttt aaggctcaga aaggtagcac
     1981 attcggataa acctggatca acctcaactg catccttcag agataatgtc accagtcctc
     2041 ttccttcact tcttgttgta attgcagcca ttttcattgg attctttcta gggaaattca
     2101 tcttgtagca attctgcagt cgacggtacc gcgggcccgg gatccaccgg atctagataa
     2161 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     2221 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     2281 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     2341 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     2401 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     2461 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     2521 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     2581 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     2641 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     2701 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     2761 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     2821 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     2881 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2941 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     3001 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3061 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3121 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3181 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3241 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     3301 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     3361 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3421 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     3481 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3541 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     3601 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     3661 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     3721 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     3781 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     3841 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     3901 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     3961 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4021 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4081 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4141 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4201 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4261 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4321 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     4381 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     4441 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     4501 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     4561 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     4621 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     4681 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     4741 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     4801 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     4861 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4921 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     4981 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5041 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5101 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5161 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     5221 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     5281 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     5341 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     5401 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     5461 aaccgtatta ccgccatgca ttagt
//
