LOCUS       sequence-479762-        5493 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of syntaxin-1(E49TAG) fused to mCherry for 
            genetic code expansion via Amber codon supression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5493)
  AUTHORS   Obashi K, Strub MP, Taraska JW
  TITLE     A PIP2-stabilized syntaxin-1a structure mapped with transition metal
            ion FRET and unnatural fluorescent amino acids at the plasma 
            membrane.
  JOURNAL   Structure. 2025 Nov 24:S0969-2126(25)00388-0. doi: 
            10.1016/j.str.2025.10.001.
  PUBMED    41290003
REFERENCE   2  (bases 1 to 5493)
  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"; journalName: "Structure. 
            2025 Nov 24:S0969-2126(25)00388-0. doi: 10.1016/j.str.2025.10.001."
COMMENT     Sequence Label: Syntaxin1A(E49TAG)-mCherry
FEATURES             Location/Qualifiers
     source          1..5493
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1495..2202
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELY"
     primer_bind     complement(1640..1658)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1908..1927
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2353..2474
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2390..2409)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2444..2463
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2481..2936)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2618..2639)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2830..2849
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2963..3067
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3092..3112)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3136..3346
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3158..3354
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3205..3340
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3389..4183
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3389..4183
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3443..3462)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4053..4072
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4371..4390)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4415..4462
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4791..5379
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5280..5299
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatgaagga ccgaacccag gagctccgca cggccaagga cagcgatgac
      661 gacgatgatg tcactgtcac tgtggaccga gaccgcttca tggatgagtt ctttgaacag
      721 gtggaagaga tccggggctt tattgacaag attgcttaga acgtggagga ggtgaagagg
      781 aaacacagcg ccatcctggc ctccccgaac cccgatgaga agaccaagga ggaactggag
      841 gagctcatgt cggacattaa gaagacagcg aacaaagttc gctccaagct aaagagcatc
      901 gagcagagca tcgagcagga ggaaggtctg aaccgctcgt cggcggacct gaggatccgg
      961 aagacgcagc attccacgct gtcccgaaag tttgtggagg tcatgtccga gtacaacgcc
     1021 actcagtcag actaccgaga acgctgcaaa gggcgcatcc agaggcagct ggagatcact
     1081 ggccggacca cgaccagtga ggagttggaa gacatgctgg agagtgggaa tcccgccatc
     1141 tttgcctctg ggatcatcat ggactccagc atctcgaagc aggccctcag tgagatcgag
     1201 accaggcaca gtgagatcat caagttggag aacagcatcc gggagctaca cgatatgttc
     1261 atggacatgg ccatgctggt ggagagccag ggggagatga ttgacaggat cgagtacaat
     1321 gtggaacacg ctgtggacta cgtggagagg gccgtgtctg acaccaagaa ggccgtcaag
     1381 taccagagca aggcacgcag gaagaagatc atgatcatca tttgctgtgt gattctgggc
     1441 atcatcatcg cctccaccat cgggggcatc tttggaggcg gatccggagg gtctatggtg
     1501 agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt caaggtgcac
     1561 atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc
     1621 tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttcgcc
     1681 tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa gcaccccgcc
     1741 gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg
     1801 aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca ggacggcgag
     1861 ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc cgtaatgcag
     1921 aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga cggcgccctg
     1981 aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga cgctgaggtc
     2041 aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa cgtcaacatc
     2101 aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta cgaacgcgcc
     2161 gagggccgcc actccaccgg cggcatggac gagctgtact gagtcgacgg taccgcgggc
     2221 ccgggatcca ccggatctag ataactgatc ataatcagcc ataccacatt tgtagaggtt
     2281 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     2341 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     2401 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     2461 atcaatgtat cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     2521 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     2581 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     2641 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     2701 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     2761 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     2821 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     2881 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg
     2941 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     3001 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     3061 gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag
     3121 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3181 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     3241 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     3301 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
     3361 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3421 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3481 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3541 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
     3601 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     3661 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     3721 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3781 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3841 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3901 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3961 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4021 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4081 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4141 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4201 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4261 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4321 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     4381 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     4441 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     4501 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     4561 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     4621 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     4681 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     4741 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     4801 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     4861 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     4921 cagataccaa atacttttct tctagtgtag ccgtagttag gccaccactt caagaactct
     4981 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     5041 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     5101 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5161 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     5221 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     5281 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     5341 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     5401 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     5461 gattctgtgg ataaccgtat taccgccatg cat
//
