LOCUS       sequence-201409-        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        71..374
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        375..578
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     529..549
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      617..626
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(623..625,626..628,629..1339)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(623..625,626..628,629..1339)
                     /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(668..689)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(929..948)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1276..1297
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2283..2404
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2320..2339)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2374..2393
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2411..2866)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2548..2569)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2760..2779
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2893..2997
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2999..3356
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2999..3190
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3022..3042)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3207..3342
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3391..4185
                     /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             3391..4185
                     /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(3445..3464)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4055..4074
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4373..4392)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4417..4464
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4793..5381
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5282..5301
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgccatgcat tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
       61 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      121 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      181 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      241 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      301 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      361 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      421 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      481 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      541 gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc
      601 gctagcgcta ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
      661 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
      721 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
      781 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
      841 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
      901 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
      961 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     1021 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     1081 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     1141 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     1201 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     1261 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     1321 catggacgag ctgtacaagt ccggactcag atctcgagct caagcttcga attgtgcgtc
     1381 cgcctcaggg gccatggcga agcacgagca gatcctggtc ctcgatccgc ccacagacct
     1441 caaattcaaa ggccccttca cagatgtagt cactacaaat cttaaattgc gaaatccatc
     1501 ggatagaaaa gtgtgtttca aagtgaagac tacagcacct cgccggtact gtgtgaggcc
     1561 caacagtgga attattgacc cagggtcaac tgtgactgtt tcagtaatgc tacagccctt
     1621 tgactatgat ccgaatgaaa agagtaaaca cgactttgac gtccagacaa tttttgctcc
     1681 accaaacact tcagatatgg aagctgtgtg gaaagaggca aaacctgatg aattaatgga
     1741 ttccaaattg agatgcgtat ttgaaatgcc caatgaaaat gataaattga atgatatgga
     1801 acctagcaaa gctgttccac tgaatgcatc taagcaagat ggacctatgc caaaaccaca
     1861 cagtgtttca cttaatgata ccgaaacaag gaaactaatg gaagagtgta aaagacttca
     1921 gggagaaatg atgaagctat cagaagaaaa tcggcacctg agagatgaag gtttaaggct
     1981 cagaaaggta gcacattcgg ataaacctgg atcaacctca actgcatcct tcagagataa
     2041 tgtcaccagt cctcttcctt cacttcttgt tgtaattgca gccattttca ttggattctt
     2101 tctagggaaa ttcatcttgt agcaattctg cagtcgacgg taccgcgggc ccgggatcca
     2161 ccggatctag ataactgatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     2221 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     2281 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     2341 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     2401 cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     2461 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     2521 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     2581 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     2641 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     2701 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     2761 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     2821 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
     2881 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     2941 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct
     3001 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     3061 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     3121 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3181 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3241 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     3301 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc
     3361 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     3421 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     3481 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     3541 accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg
     3601 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     3661 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     3721 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     3781 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     3841 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     3901 ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     3961 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     4021 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     4081 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     4141 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     4201 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     4261 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     4321 agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac
     4381 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     4441 cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
     4501 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
     4561 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
     4621 ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat
     4681 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     4741 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4801 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4861 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4921 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4981 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     5041 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     5101 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     5161 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     5221 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     5281 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     5341 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     5401 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     5461 ctgattctgt ggataaccgt attac
//
