LOCUS       sequence-384268-        6803 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid encoding SUMO-tagged full-length Syt9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6803)
  AUTHORS   Seibert MJ, Evans CS, Stanley KS, Wu Z, Chapman ER
  TITLE     Synaptotagmin 9 Modulates Spontaneous Neurotransmitter Release in 
            Striatal Neurons by Regulating Substance P Secretion.
  JOURNAL   J Neurosci. 2023 Mar 1;43(9):1475-1491. doi: 
            10.1523/JNEUROSCI.1857-22.2023. Epub 2023 Feb 2.
  PUBMED    36732068
REFERENCE   2  (bases 1 to 6803)
  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: 
            "10.1523/JNEUROSCI.1857-22.2023"; journalName: "J Neurosci"; date: 
            "2023-03-1- 1"; volume: "43"; issue: "9"; pages: "1475-1491"
COMMENT     Sequence Label: pET (SUMO)-Syt9
FEATURES             Location/Qualifiers
     source          1..6803
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     158..177
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     261..280
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        261..279
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    280..304
                     /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)."
     RBS             319..341
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             361..378
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             388..405
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             412..705
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             1928..1945
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1998..2016)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2012..2059
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2096..2551
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2183..2202)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2393..2414
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2643..3458)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2643..3458)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3366..3385
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3450..3469
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3580..4168
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4069..4088
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4322..4339
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4354..4493
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4419..4438)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4579..4601
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4595..4786)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5595..6677)
                     /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     6639..6658
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6678..6755)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga
       61 tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc
      121 tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc
      181 cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatcg
      241 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
      301 ttcccctcta gaaataattt tgtttaactt taagaaggag atatacatat gggcagcagc
      361 catcatcatc atcatcacgg cagcggcctg gtgccgcgcg gcagcgctag catgtcggac
      421 tcagaagtca atcaagaagc taagccagag gtcaagccag aagtcaagcc tgagactcac
      481 atcaatttaa aggtgtccga tggatcttca gagatcttct tcaagatcaa aaagaccact
      541 cctttaagaa ggctgatgga agcgttcgct aaaagacagg gtaaggaaat ggactcctta
      601 agattcttgt acgacggtat tagaattcaa gctgatcaga cccctgaaga tttggacatg
      661 gaggataacg atattattga ggctcacaga gaacagattg gtggatccat gttcccggaa
      721 cccccgaccc cggggtctcc agcgcctgag acacctccag actccagccg catcaggcag
      781 ggggcagtgc ctgcctgggt cctggctacc atcttgctgg gctcaggcct cctggtcttc
      841 agcagctgtt tctgtctcta ccggaagcgc tgtcggagac ggatgggcaa aaagagtcag
      901 gcccaagccc aagtccatct tcaggaagtg aaggagctgg gccggagtta catagataag
      961 gttcagcctg aaatagagga actggacccc tcaccatcca tgccaggaca gcaggtattg
     1021 gacaagcacc agttaggccg actgcagtat tcactggatt atgacttcca gactggccag
     1081 ctcctggtag gcatcttgca agctgaggga ctggcagcgt tggacctagg aggttcctca
     1141 gacccctatg ttagtgtcta tctgctgcca gacaagcgga ggcgacatga gaccaaggtg
     1201 catcggcaga ccctgaatcc acactttgga gagacctttg ccttcaaggt cccatacgtg
     1261 gaactaggag gtagagtgct ggtcatggcg gtgtatgact tcgatcgctt ctcccgcaac
     1321 gatgccatcg gagaggtgcg ggtgcctatg agttcagtga acctggggcg gccagtgcag
     1381 gcctggcgag agctgcaggt ggctcccaaa gaggagcagg agaaactggg ggatatctgc
     1441 ttctctctcc ggtatgtccc cacggccggg aagctcaccg tcattgtcct ggaagctaaa
     1501 aacctgaaga aaatggatgt aggaggactc tcagaccctt atgtcaaggt gcacctgctc
     1561 cagggaggta aaaaggttcg gaagaagaaa accaccatta agaagaacac cctgaacccc
     1621 tattacaacg aggccttcag ctttgaggtg ccctgtgacc aggtgcagaa agtccaggtg
     1681 gagctgaccg ttttggacta tgacaaactg gggaagaatg aggccatcgg gagagtggcg
     1741 gtgggcacag cagttggtgg ggctggccta cggcactggg ctgacatgtt ggccaaccct
     1801 aggcggccca ttgcccagtg gcactctctg cggccccctg acagagccag gccaatacct
     1861 gcaccctgaa agcttaggta tttattcggc gcaaagtgcg tcgggtgatg ctgccaactt
     1921 agtcgagcac caccaccacc atcactgaga tccggctgct aacaaagccc gaaaggaagc
     1981 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     2041 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg
     2101 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2161 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2221 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2281 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2341 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2401 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2461 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2521 aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg
     2581 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgaattaat
     2641 tcttagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca
     2701 ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc
     2761 cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa
     2821 cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg
     2881 actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg ttcaacaggc
     2941 cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat
     3001 tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc
     3061 gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga
     3121 tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag taaccatgca
     3181 tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag
     3241 tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga
     3301 aacaactctg gcgcatcggg cttcccatac aatcgataga ttgtcgcacc tgattgcccg
     3361 acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc
     3421 ggcctagagc aagacgtttc ccgttgaata tggctcataa caccccttgt attactgttt
     3481 atgtaagcag acagttttat tgttcatgac caaaatccct taacgtgagt tttcgttcca
     3541 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3601 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3661 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3721 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3781 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3841 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3901 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3961 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4021 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4081 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4141 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4201 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     4261 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4321 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     4381 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     4441 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     4501 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4561 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4621 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     4681 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     4741 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     4801 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     4861 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     4921 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     4981 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     5041 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     5101 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     5161 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     5221 tcaacgacag gagcacgatc atgcgcaccc gtggggccgc catgccggcg ataatggcct
     5281 gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca
     5341 agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga aagcggtcct
     5401 cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg ataaagaaga
     5461 cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag ctgactgggt
     5521 tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc taacttacat
     5581 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5641 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt
     5701 cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc ctgagagagt
     5761 tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt
     5821 aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac cgagatatcc
     5881 gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg
     5941 ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga
     6001 aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga
     6061 gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc
     6121 gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta
     6181 ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat
     6241 aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga
     6301 tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag
     6361 gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg
     6421 cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca
     6481 acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa
     6541 ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc
     6601 tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat
     6661 aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg ctatcatgcc
     6721 ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct ctcccttatg
     6781 cgactcctgc attaggaagc agc
//
