LOCUS       sequence-192821-        6032 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human Stx3 locked into the open conformation with a C-term
            myc-myc-his tag in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6032)
  AUTHORS   Giovannone AJ, Reales E, Bhattaram P, Fraile-Ramos A, Weimbs T
  TITLE     Monoubiquitination of syntaxin 3 leads to retrieval from the 
            basolateral plasma membrane and facilitates cargo recruitment to 
            exosomes.
  JOURNAL   Mol Biol Cell. 2017 Oct 15;28(21):2843-2853. doi: 
            10.1091/mbc.E17-07-0461. Epub 2017 Aug 16.
  PUBMED    28814500
REFERENCE   2  (bases 1 to 6032)
  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: 
            "10.1091/mbc.E17-07-0461"; journalName: "Mol Biol Cell"; date: 
            "2017-10-15- 15"; volume: "28"; issue: "21"; pages: "2843-2853"
COMMENT     Sequence Label: pcDNA4/TO h-Stx3 L165A/E166A-2xMyc/His
FEATURES             Location/Qualifiers
     source          1..6032
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    695..713
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    716..734
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     744..768
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1491..1502
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             1791..1820
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1830..1859
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1875..1892
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1915..1932)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1921..2145
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2191..2619
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2278..2297)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2488..2509
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2628..2648)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2633..2962
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2813..2948
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2875..2894
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3010..3057
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3076..3450
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    3580..3701
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3617..3636)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3671..3690
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3750..3766)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3750..3766)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3763..3785)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3774..3790
                     /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)."
     promoter        complement(join(3798..3804,3805..3822,3823..3828))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3843..3864
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3981..3998)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4152..4740)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4232..4251)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4911..5702,5703..5771))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5534..5553
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5772..5876)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5951..5970)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      361 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      421 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      481 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      601 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      661 cgtgtacggt gggaggtcta tataagcaga gctctcccta tcagtgatag agatctccct
      721 atcagtgata gagatcgtcg acgagctcgt ttagtgaacc gtcagatcgc ctggagacgc
      781 catccacgct gttttgacct ccatagaaga caccgggacc gatccagcct ccggactcta
      841 gcgtttaaac ttaagcttgg taccgagctc ggatccacta gtccagtgtg gtggaattcc
      901 tgggcttcag gatgaaggac cgtctggagc agctgaaggc caagcagctg acacaggatg
      961 atgatactga tgcggttgag attgctatcg acaacacggc ttttatggac gagttctttt
     1021 ctgagattga ggaaactcgg cttaacattg acaagatctc agaacatgta gaggaggcta
     1081 agaaactcta cagtatcatt ctctctgcac cgattccaga gccaaaaacc aaggatgacc
     1141 tagagcagct cacgactgag attaagaaaa gggccaacaa cgtccggaac aaactgaaga
     1201 gcatggagaa gcatattgaa gaagatgagg tcaggtcatc ggcagacctt cggattcgga
     1261 aatcccagca ctctgtcctt tctcggaagt ttgtggaggt gatgaccaaa tacaatgaag
     1321 ctcaagtgga cttccgagaa cgcagcaaag ggcgaatcca gcggcagctc gaaattactg
     1381 gcaaaaagac aaccgatgag gaggcggcgg agatgttgga gagtggcaac ccggccatct
     1441 tcacttctgg gatcattgac tcacagattt ccaagcaagc cctcagtgag attgagggac
     1501 gacacaagga cattgtgagg ctggagagca gcatcaagga gcttcacgac atgtttatgg
     1561 acatcgccat gctggtggag aatcagggtg agatgttaga taacatagag ttgaatgtca
     1621 tgcacacagt ggaccacgtg gagaaggcac gagatgaaac gaaaaaagct gtgaaatacc
     1681 agagtcaggc ccggaagaaa ttgataatta tcattgtgct agtagttgtg ttgctgggca
     1741 ttttagcatt gattattgga ctttccgttg ggctgaatgc ggccgctcga gaacaaaaac
     1801 tcatctcaga agaggatctg gggcccttcg aacaaaaact catctcagaa gaggatctga
     1861 atatgcatac cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga
     1921 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     1981 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2041 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2101 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2161 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     2221 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2281 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2341 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2401 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2461 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2521 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2581 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2641 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2701 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2761 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     2821 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     2881 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     2941 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3001 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     3061 gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
     3121 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
     3181 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
     3241 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
     3301 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
     3361 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
     3421 gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
     3481 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3541 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3601 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3661 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3721 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3781 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3841 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3901 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3961 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4021 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4081 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4141 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4201 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4261 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4321 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4381 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4441 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4501 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4561 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4621 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4681 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4741 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4801 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4861 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4921 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4981 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5041 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5101 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5161 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5221 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5281 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5341 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5401 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5461 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5521 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5581 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5641 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5701 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5761 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5821 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5881 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
     5941 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
     6001 gtgtgttgga ggtcgctgag tagtgcgcga gc
//
