LOCUS       sequence-391935-        6661 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Synaptotagmin 3 with pH-sensitive 
            fluorescent protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6661)
  AUTHORS   Dean C, Dunning FM, Liu H, Bomba-Warczak E, Martens H, Bharat V, 
            Ahmed S, Chapman ER
  TITLE     Axonal and dendritic synaptotagmin isoforms revealed by a 
            pHluorin-syt functional screen.
  JOURNAL   Mol Biol Cell. 2012 May;23(9):1715-27. doi: 10.1091/mbc.E11-08-0707.
            Epub 2012 Mar 7.
  PUBMED    22398727
REFERENCE   2  (bases 1 to 6661)
  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"; doi: 
            "10.1091/mbc.E11-08-0707"; journalName: "Mol Biol Cell"; date: 
            "2012-05"; volume: "23"; issue: "9"; pages: "1715-27"
COMMENT     Sequence Label: pCI pHluorin Syt3
FEATURES             Location/Qualifiers
     source          1..6661
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     718..742
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1034..1052
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     5'UTR           1076..1118
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     CDS             1127..1216
                     /codon_start=1
                     /product="signal sequence from bovine prolactin"
                     /label=prolactin signal sequence
                     /translation="MDSKGSSQKGSRLLLLLVVSNLLLCQGVVS"
     CDS             1235..1945
                     /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="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIKAN
                     FKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(1262..1290)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1885..1905
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    3775..3896
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3786..3805)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3840..3859
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      4077..4532
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4164..4183)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4374..4395
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4599..4618)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4718..4740
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4778..4796)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4864..4968
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4969..5037,5038..5829)
                     /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(5187..5206)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6000..6588
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6489..6508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tcgtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt acttaatacg actcactata ggctagcctc gagaattcga tatcgcttgt
     1081 tctttttgca gaagctcaga ataaacgctc aactttggca gataccatgg acagcaaagg
     1141 ttcgtcgcag aaagggtccc gcctgctcct gctgctggtg gtgtcaaatc tactcttgtg
     1201 ccagggtgtg gtctccgata tcggtaccac gcgtagtaaa ggcgaagaac ttttcactgg
     1261 agttgtccca attcttgttg aattagatgg tgatgttaat gggcacaaat tttctgtcag
     1321 tggagagggt gaaggtgatg caacatacgg aaaacttacc cttaaattta tttgcactac
     1381 tggaaaacta cctgttcctt ggccaacact tgtcactact ttaacttatg gtgttcaatg
     1441 cttttcaaga tacccagatc atatgaaacg gcatgacttt ttcaagagtg ccatgcccga
     1501 aggttatgta caggaaagaa ctatattttt caaagatgac gggaactaca agacacgtgc
     1561 tgaagtcaag tttgaaggtg atacccttgt taatagaatc gagttaaaag gtattgattt
     1621 taaagaagat ggaaacattc ttggacacaa attggaatac aactataacg atcaccaggt
     1681 gtacatcatg gcagacaaac aaaagaatgg aatcaaagct aacttcaaaa ttagacacaa
     1741 cattgaagat ggaggcgttc aactagcaga ccattatcaa caaaatactc caattggcga
     1801 tgggcccgtc cttttaccag acaaccatta cctgtttaca acttctactc tttcgaaaga
     1861 tcccaacgaa aagagagacc acatggtcct tcttgagttt gtaacagctg ctgggattac
     1921 acatggcatg gatgaactat acaaagaatt ctcaggcgga agcggaggct ccggcgggag
     1981 cggcggatct ggggactatg aagatgacct ctgtcggcgg gccctcatcc tggtttcaga
     2041 cctctgtgca cggatccgag atgctgatac caatgacagg tgtcaagagt tcaacgaact
     2101 gcgaatcaga ggatatcccc ggggtccgga tgcagacatt tccgtgagcc tgctgtcagt
     2161 catcgtgacg ttctgtggca tcgtcctctt gggtgtctct ctcttcgtgt cctggaagtt
     2221 gtgctgggtg ccctggaggg acaaaggggg ctcagcagtg ggtggtggcc ccctgcgcaa
     2281 agacttagcc cctggggttg ggctggcagg cctggtagga ggtggtgggc accatctggg
     2341 agccagccta ggtggccatc ccctgctggg tggtccgcac catcatgcac acccagccca
     2401 ccatccacca tttgctgaac tgctggaacc aggtggcctt gggggctctg agcctccaga
     2461 accttcctac ctggacatgg actcatatcc agaggctgct gtggcctctg tggtagctgc
     2521 tggggtcaaa ccaagccaga catcccctga gctgccttct gagggtggaa caggctctgg
     2581 gctgctcctg ctgcctccca gtggtggggg cttgcccagt gcccagtctc atcagcaggt
     2641 cacaagtcta gcacccacca ccaggtaccc agccctgcct cggcccctca cccagcagac
     2701 cctgaccacc caagcagacc caagcagtga ggagcgacca cctgcactgc ccttacccct
     2761 gccaggcgga gaggagaaag ccaagctcat cggtcagatc aagccagagc tataccaggg
     2821 cactggacct ggtggccggc gaagcggcgg gggctctggg gaggcagggg caccctgtgg
     2881 ccgcatcagt tttgcccttc ggtacctcta tggctctgac cagctcgtgg tgcggatcct
     2941 tcaggccctg gatctcccag caaaggactc caatgggttc tcagacccct acgtcaagat
     3001 ctacctgttg cctgatcgca aaaagaagtt ccagaccaag gtgcacagga agacgctgaa
     3061 ccctatcttc aacgagacgt ttcagttttc ggtacccttg gccgagctgg cgcagcgcaa
     3121 gctgcacttc agtgtctatg acttcgacag attctcgagg cacgatctca tcggccaggt
     3181 ggttttggac aacctgctgg agcttgctga acagccccct gaccgtccac tctggaggga
     3241 catcctggag ggtggctcgg aaaaggcaga tcttggggag ctcaacttct cactctgcta
     3301 cctccccacg gccgggcgcc tcaccgtgac catcatcaaa gcctctaacc tcaaagcaat
     3361 ggacctcact ggtttctcag acccctacgt gaaggcctct ctgatcagcg aggggcggcg
     3421 tctgaagaaa cggaagacct ccattaagaa gaacacgctg aaccccacat acaacgaggc
     3481 cctggtgttc gacgtggctc cggagagcgt ggagaacgtg ggtctcagta tcgcagtggt
     3541 agactatgac tgcatcggcc acaatgaagt gatcggggtg tgccgcgtgg gcccagaggc
     3601 cgctgaccca catggcagag aacactgggc agagatgctg gctaaccctc gcaagcctgt
     3661 ggagcactgg caccagctag tggaggagaa gactctaagc agctttacca agggcggcaa
     3721 aggattgtca gagaaagaga attcagagtg agcggccgct tcgagcagac atgataagat
     3781 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     3841 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     3901 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa
     3961 gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tccgggctgg cgtaatagcg
     4021 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggacgc
     4081 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4141 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     4201 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     4261 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     4321 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     4381 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     4441 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     4501 gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat
     4561 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     4621 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     4681 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     4741 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     4801 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     4861 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     4921 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     4981 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     5041 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     5101 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     5161 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     5221 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     5281 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     5341 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     5401 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     5461 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     5521 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     5581 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     5641 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     5701 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     5761 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     5821 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     5881 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     5941 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     6001 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     6061 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     6121 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     6181 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     6241 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     6301 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     6361 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     6421 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     6481 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     6541 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     6601 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatggc tcgacagatc
     6661 t
//
