LOCUS       sequence-317164-        5815 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes cytoplasmic domain of synaptotagmin-1 for bacterial 
            expression, purification, and labeling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5815)
  AUTHORS   Bradberry MM, Bao H, Lou X, Chapman ER
  TITLE     Phosphatidylinositol 4,5-bisphosphate drives Ca(2+)-independent 
            membrane penetration by the tandem C2 domain proteins 
            synaptotagmin-1 and Doc2beta.
  JOURNAL   J Biol Chem. 2019 Jul 12;294(28):10942-10953. doi: 
            10.1074/jbc.RA119.007929. Epub 2019 May 30.
  PUBMED    31147445
REFERENCE   2  (bases 1 to 5815)
  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: 
            "10.1074/jbc.RA119.007929"; journalName: "J Biol Chem"; date: 
            "2019-07-12- 12"; volume: "294"; issue: "28"; pages: "10942-10953"
COMMENT     Sequence Label: pGEX-4T Syt1 C2AB(140) M173C
FEATURES             Location/Qualifiers
     source          1..5815
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..22,23..29)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    37..53
                     /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)."
     CDS             76..729
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     687..709
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             736..753
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1683..1705)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1850..1868)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1936..2040
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2041..2109,2110..2901)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2259..2278)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3072..3660
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3561..3580
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3814..3831
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3904..3981
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3982..5064
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(4001..4020)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5113..5118,5119..5136,5137..5143)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5151..5167
                     /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     5175..5191
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5175..5191
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5187..5714
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5207..5224)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5207..5223)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5216..5238)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttgacaatta atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca
       61 caggaaacag tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      121 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      181 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      241 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      301 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      361 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      421 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      481 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
      541 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
      601 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
      661 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
      721 cctccaaaat cggatctggt tccgcgtgga tccggatctg gatctgagaa actgggaaag
      781 ctccaatatt cactggacta tgacttccag aataaccagc tgttggtggg aatcatccag
      841 gctgctgaac tgcccgccct ggactgcggg ggtacatccg atccatacgt caaagtcttc
      901 ctgctgcctg acaaaaagaa gaaatttgag actaaagtcc accggaaaac cctcaatcca
      961 gtcttcaatg aacaatttac tttcaaggta ccctactcgg aattaggtgg caaaaccctg
     1021 gtgatggctg tgtatgactt tgatcgcttc tccaagcacg acatcatcgg agagttcaaa
     1081 gttcctatga acaccgtgga ttttggccat gtgaccgagg agtggcgcga tctccagagc
     1141 gctgagaaag aagagcaaga gaaactgggt gacatcgcct tctccctccg ctacgtccct
     1201 actgccggca aactgactgt tgtcattctg gaagccaaga acctgaagaa gatggatgtg
     1261 ggtggcttat ctgatcccta cgtgaagatt cacctgatgc agaacggtaa gaggctgaag
     1321 aagaaaaaga cgacgattaa gaagaacaca ctcaacccct actacaacga gtccttcagc
     1381 tttgaagttc cgttcgagca aatccagaaa gtgcaagtgg tggtaactgt tttggactat
     1441 gacaagattg gcaagaacga cgccatcgga aaagtcttcg ttggttacaa cagcactggg
     1501 gcggagctgc gacactggtc agacatgctg gccaaccccc ggcgacccat cgcacagtgg
     1561 cacactctgc aggtagagga ggaggttgat gccatgctgg ctgtcaagaa gtaagaattc
     1621 atcgggccgc atcgtgactg actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa
     1681 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     1741 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     1801 acccagtcac gtagcgatag cggagtgtat aattcttgaa gacgaaaggg cctcgtgata
     1861 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     1921 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     1981 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     2041 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     2101 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     2161 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     2221 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     2281 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     2341 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     2401 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2461 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2521 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2581 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2641 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2701 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     2761 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     2821 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     2881 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     2941 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     3001 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     3061 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     3121 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     3181 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     3241 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3301 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3361 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3421 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3481 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3541 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3601 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3661 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3721 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3781 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     3841 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
     3901 tccgacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
     3961 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
     4021 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
     4081 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
     4141 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
     4201 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
     4261 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
     4321 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
     4381 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
     4441 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
     4501 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
     4561 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
     4621 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     4681 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     4741 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     4801 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     4861 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     4921 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     4981 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     5041 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     5101 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     5161 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     5221 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     5281 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     5341 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     5401 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     5461 agatgcacgg ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc
     5521 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     5581 aaagctggct acaggaaggc cagacgcgaa ttatttttga tggcgttgga attagcttat
     5641 cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc
     5701 tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat
     5761 aatgtttttt gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctg
//
