LOCUS       sequence-342069-        5823 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Reporter for VAMP4 exocytosis in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5823)
  TITLE     The vSNAREs VAMP2 and VAMP4 control recycling and intracellular 
            sorting of post-synaptic receptors in neuronal dendrites
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5823)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: VAMP4-SEP
FEATURES             Location/Qualifiers
     source          1..5823
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..46)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(29..45)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(38..60)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      258..713
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(345..364)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     555..576
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(730..749)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     849..871
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(909..927)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        995..1099
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1100..1168,1169..1960)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1318..1337)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2131..2719
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2620..2639
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2873..2890
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3007..3028
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3043..3048,3049..3066,3067..3073)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3081..3097
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     3086..3108
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3105..3121
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3105..3121
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        3150..3529
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3530..3733
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3684..3704
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3730..3754
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        3964..3982
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3964..3981
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     3995..4014
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4511..5224
                     /codon_start=1
                     /product="pH-sensitive mutant of green fluorescent protein 
                     (Ng et al., 2002)"
                     /label=superecliptic pHluorin
                     /note="excited strongly by blue light at neutral pH but 
                     minimally fluorescent at acidic pH"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(4541..4569)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     5164..5184
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     regulatory      5259..5268
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    5272..5406
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5309..5328)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5363..5382
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        join(5475..5823,1..9)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5475..5666
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5498..5518)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5683..5818
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 ttttgcaaaa agctgttaac agcttggcac tggccgtcgt tttacaacgt cgtgactggg
       61 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
      121 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
      181 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
      241 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
      301 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
      361 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
      421 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
      481 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
      541 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggg
      601 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
      661 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
      721 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
      781 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
      841 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
      901 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
      961 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     1021 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     1081 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     1141 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     1201 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     1261 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     1321 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     1381 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     1441 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     1501 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     1561 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     1621 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     1681 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     1741 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     1801 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     1861 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     1921 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     1981 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     2041 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     2101 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     2161 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     2221 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     2281 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     2341 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     2401 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     2461 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     2521 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     2581 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     2641 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     2701 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     2761 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     2821 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     2881 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     2941 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     3001 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     3061 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     3121 catgattacg aattaattcg agctcgcccg acattgatta ttgactagtt attaatagta
     3181 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
     3241 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
     3301 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
     3361 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
     3421 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
     3481 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
     3541 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
     3601 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
     3661 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
     3721 tataagcaga gctcgtttag tgaaccgtca gatcgcctgg agacgccatc cacgctgttt
     3781 tgacctccat agaagacacc gggaccgatc cagcctccgc ggccgggaac ggtgcattgg
     3841 aacgcggatt ccccgtgcca agagtgacgt aagtaccgcc tatagagtct ataggcccac
     3901 ccccttggct tcgttagaac gcggctacaa ttaatacata accttatgta tcatacacat
     3961 acgatttagg tgacactata gaataacatc cactttgcct ttctctccac aggtgtccac
     4021 tcccaggtcc aactgcacct cggttctatc gattgaattc gccaccatgc ctcccaagtt
     4081 taagcgccac ctcaatgatg atgatgtcac aggttctgtg aaaagtgaaa ggagaaatct
     4141 tttggaagat gattcagatg aagaagagga cttttttcta aggggaccat ctggaccaag
     4201 atttggacct agaaatgata aaattaagca tgttcagaat caagtggatg aagttattga
     4261 tgtcatgcaa gaaaatatta caaaggtaat tgagagaggg gagagactag atgaactaca
     4321 ggacaaatca gaaagcttat cggataatgc aacagctttt agcaacagat ccaaacaact
     4381 tcgaaggcaa atgtggtggc gtggatgcaa aataaaagcc atcatggctt tggttgctgc
     4441 tatccttttg ctagtgatta tcattcttat agtcatgaaa taccgtactt cggatccacc
     4501 ggtcgccacc atgagtaaag gagaagaact tttcactgga gttgtcccaa ttcttgttga
     4561 attagatggt gatgttaatg ggcacaaatt ttctgtcagt ggagagggtg aaggtgatgc
     4621 aacatacgga aaacttaccc ttaaatttat ttgcactact ggaaaactac ctgttccttg
     4681 gccaacactt gtcactactt taacttatgg tgttcaatgc ttttcaagat acccagatca
     4741 tatgaaacgg catgactttt tcaagagtgc catgcccgaa ggttatgtac aggaaagaac
     4801 tatatttttc aaagatgacg ggaactacaa gacacgtgct gaagtcaagt ttgaaggtga
     4861 tacccttgtt aatagaatcg agttaaaagg tattgatttt aaagaagatg gaaacattct
     4921 tggacacaaa ttggaataca actataacga tcaccaggtg tacatcatgg cagacaaaca
     4981 aaagaatgga atcaaagcta acttcaaaat tagacacaac attgaagatg gaggcgttca
     5041 actagcagac cattatcaac aaaatactcc aattggcgat gggcccgtcc ttttaccaga
     5101 caaccattac ctgtttacaa cttctactct ttcgaaagat cccaacgaaa agagagacca
     5161 catggtcctt cttgagtttg taacagctgc tgggattaca catggcatgg atgaactata
     5221 caaagctagc taatctagag tcgacctgca gaagcttggc cgccatggcc caacttgttt
     5281 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     5341 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     5401 tggatcggga attaattcgg cgcagcacca tggcctgaaa taacctctga aagaggaact
     5461 tggttaggta ccttctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt
     5521 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     5581 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     5641 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     5701 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     5761 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     5821 ggc
//
