LOCUS       sequence-368935-        5627 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SNAP-23 in E. Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5627)
  AUTHORS   Lai Y, Fois G, Flores JR, Tuvim MJ, Zhou Q, Yang K, Leitz J, Peters 
            J, Zhang Y, Pfuetzner RA, Esquivies L, Jones P, Frick M, Dickey BF, 
            Brunger AT
  TITLE     Inhibition of calcium-triggered secretion by hydrocarbon-stapled 
            peptides.
  JOURNAL   Nature. 2022 Mar;603(7903):949-956. doi: 10.1038/s41586-022-04543-1.
            Epub 2022 Mar 23.
  PUBMED    35322233
REFERENCE   2  (bases 1 to 5627)
  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: 
            "10.1038/s41586-022-04543-1"; journalName: "Nature"; date: 
            "2022-03"; volume: "603"; issue: "7903"; pages: "949-956"
COMMENT     Sequence Label: SNAP-23
FEATURES             Location/Qualifiers
     source          1..5627
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(202..207,208..225,226..232)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    240..256
                     /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     264..280
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     264..280
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             276..803
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(296..313)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(296..312)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(305..327)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(905..910,911..926,927..933)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    941..957
                     /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             980..1633
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1591..1613
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1640..1657
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2399..2421)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2566..2584)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2652..2756
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2757..2825,2826..3617)
                     /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(2975..2994)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3788..4376
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4277..4296
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4530..4547
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4620..4697
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4698..5627,1..153)
                     /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(4717..4736)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg
       61 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      121 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct
      181 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      241 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg gattcactgg
      301 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
      361 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
      421 cccaacagtt gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg gcaccagaag
      481 cggtgccgga aagctggctg gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct
      541 caaactggca gatgcacggt tacgatgcgc ccatctacac caacgtgacc tatcccatta
      601 cggtcaatcc gccgtttgtt cccacggaga atccgacggg ttgttactcg ctcacattta
      661 atgttgatga aagctggcta caggaaggcc agacgcgaat tatttttgat ggcgttggaa
      721 ttagcttatc gactgcacgg tgcaccaatg cttctggcgt caggcagcca tcggaagctg
      781 tggtatggct gtgcaggtcg taaatcactg cataattcgt gtcgctcaag gcgcactccc
      841 gttctggata atgttttttg cgccgacatc ataacggttc tggcaaatat tctgaaatga
      901 gctgttgaca attaatcatc ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt
      961 cacacaggaa acagtattca tgtcccctat actaggttat tggaaaatta agggccttgt
     1021 gcaacccact cgacttcttt tggaatatct tgaagaaaaa tatgaagagc atttgtatga
     1081 gcgcgatgaa ggtgataaat ggcgaaacaa aaagtttgaa ttgggtttgg agtttcccaa
     1141 tcttccttat tatattgatg gtgatgttaa attaacacag tctatggcca tcatacgtta
     1201 tatagctgac aagcacaaca tgttgggtgg ttgtccaaaa gagcgtgcag agatttcaat
     1261 gcttgaagga gcggttttgg atattagata cggtgtttcg agaattgcat atagtaaaga
     1321 ctttgaaact ctcaaagttg attttcttag caagctacct gaaatgctga aaatgttcga
     1381 agatcgttta tgtcataaaa catatttaaa tggtgatcat gtaacccatc ctgacttcat
     1441 gttgtatgac gctcttgatg ttgttttata catggaccca atgtgcctgg atgcgttccc
     1501 aaaattagtt tgttttaaaa aacgtattga agctatccca caaattgata agtacttgaa
     1561 atccagcaag tatatagcat ggcctttgca gggctggcaa gccacgtttg gtggtggcga
     1621 ccatcctcca aaatcggatc tggttccgcg tggatccccg ggaatttccg gtggtggtgg
     1681 tggaattctg atggataatc tgtcatcaga agaaattcaa cagagagctc accagattac
     1741 tgatgagtct ctggaaagta cgaggagaat cctgggttta gccattgagt ctcaggatgc
     1801 aggaatcaag accatcacta tgctggatga acaaaaggaa caactaaacc gcatagaaga
     1861 aggcttggac caaataaata aggacatgag agagacagag aagactttaa cagaactcaa
     1921 caaaagcagt ggccttagtg tcagcccaag taatagaaca aagaactttg agtctggcaa
     1981 ggcttataag acaacatggg gagatggtgg agaaaactca cctagcaatg tagtatctaa
     2041 acagccaggc ccggtgacaa atggtcagct tcagcaacca acaacaggag cagtcagtgg
     2101 tggatacatt aaacgcataa ctaatgatgc cagagaagat gaaatggaag agaacctgac
     2161 tcaagtgggc agtatcctgg gaaatctaaa agacatggcc ctgaacatag gcaatgagat
     2221 tgatgctcaa aatccacaaa taaaacgaat cacagacaag gctgacacca acagagatcg
     2281 tattgatatt gccaatgcca gagcaaagaa actcattgac agctaactcg agctcaagct
     2341 taattcatcg tgactgactg acgatctgcc tcgcgcgttt cggtgatgac ggtgaaaacc
     2401 tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca
     2461 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc
     2521 agtcacgtag cgatagcgga gtgtataatt cttgaagacg aaagggcctc gtgatacgcc
     2581 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     2641 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2701 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     2761 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     2821 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     2881 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     2941 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     3001 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     3061 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     3121 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     3181 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3241 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3301 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3361 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3421 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3481 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3541 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3601 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     3661 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     3721 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3781 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     3841 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3901 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     3961 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     4021 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     4081 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     4141 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     4201 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     4261 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     4321 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     4381 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     4441 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     4501 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     4561 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc ataaattccg
     4621 acaccatcga atggcgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
     4681 aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
     4741 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
     4801 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
     4861 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
     4921 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
     4981 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
     5041 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
     5101 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
     5161 ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg
     5221 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
     5281 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
     5341 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
     5401 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
     5461 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
     5521 ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     5581 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggc
//
