LOCUS       sequence-381549-        5010 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses S48A phosphodead VAMP3 fused to mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5010)
  AUTHORS   Chen T, Psoma A, Mahajan S, Ter Beest M, Linders P, Franciosa G, 
            Bianchi F, Bogaart GVD, Warner H
  TITLE     Phosphorylation of VAMP3 couples IL-6 Exocytosis to dendritic cell 
            activation.
  JOURNAL   J Cell Sci. 2025 Sep 22:jcs.264139. doi: 10.1242/jcs.264139.
  PUBMED    40977280
REFERENCE   2  (bases 1 to 5010)
  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.
            2025 Sep 22:jcs.264139. doi: 10.1242/jcs.264139."
COMMENT     Sequence Label: VAMP3(S48A)-mCherry
FEATURES             Location/Qualifiers
     source          1..5010
                     /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"
     regulatory      959..968
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             965..1675
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1110..1128)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1378..1397
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1798..1919
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1835..1854)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1889..1908
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1926..2381)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2063..2084)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2275..2294
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2408..2512
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2514..2871
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2514..2705
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2537..2557)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2722..2857
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2906..3700
                     /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             2906..3700
                     /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(2960..2979)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3570..3589
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3888..3907)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3932..3979
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4308..4896
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4797..4816
                     /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 gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attcatgtct acaggggtgc cttcagggtc
      661 cagtgctgcc actggcagta atcgaagact ccagcagaca cagaatcaag tagatgaggt
      721 ggtggacatc atgagagtca atgtggataa ggtgttagaa agagaccaga agctcgccga
      781 gctagacgac cgcgcagatg cactgcaggc aggcgcctcg cagtttgaaa caagtgctgc
      841 caagttgaag agaaagtatt ggtggaagaa ctgcaagatg tgggcgatag ggatcagtgt
      901 cctggtgatt attgtcatta tcatcattgt gtggtgtgtc tctaaggatc caccggtcgc
      961 caccatggtg agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt
     1021 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
     1081 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct
     1141 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
     1201 gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga
     1261 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
     1321 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
     1381 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
     1441 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
     1501 cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
     1561 cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta
     1621 cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agtaaagcgg
     1681 ccgcgactct agatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
     1741 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     1801 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     1861 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
     1921 ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     1981 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     2041 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     2101 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     2161 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     2221 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     2281 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     2341 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg
     2401 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2461 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc
     2521 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     2581 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     2641 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     2701 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     2761 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     2821 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca
     2881 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     2941 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     3001 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     3061 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     3121 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     3181 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     3241 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     3301 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     3361 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     3421 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     3481 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     3541 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     3601 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     3661 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     3721 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     3781 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     3841 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     3901 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     3961 ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat
     4021 accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac
     4081 cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc
     4141 atagcctcag gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     4201 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     4261 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     4321 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     4381 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     4441 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     4501 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     4561 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     4621 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     4681 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4741 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4801 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     4861 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     4921 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     4981 tctgtggata accgtattac cgccatgcat
//
