LOCUS       sequence-481089-        5565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of syntaxin-1(E53TAG,G42H,K46H) 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 5565)
  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 5565)
  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(E53TAG,G42H,K46H)-mCherry
FEATURES             Location/Qualifiers
     source          1..5565
                     /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
     promoter        3069..3426
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3069..3260
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3092..3112)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3277..3412
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3461..4255
                     /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             3461..4255
                     /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(3515..3534)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4125..4144
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4443..4462)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4487..4534
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4863..5451
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5352..5371
                     /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 tccggcactt tattgaccac attgctgaaa acgtggagta 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 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3241 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3301 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     3361 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     3421 tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     3481 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     3541 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     3601 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct
     3661 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     3721 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     3781 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     3841 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     3901 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     3961 agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac
     4021 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4081 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4141 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     4201 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     4261 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     4321 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4381 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta
     4441 actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga
     4501 cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc
     4561 tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc
     4621 ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg
     4681 gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt
     4741 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     4801 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     4861 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     4921 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     4981 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     5041 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     5101 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     5161 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     5221 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     5281 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     5341 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     5401 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     5461 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     5521 gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcat
//
