LOCUS       sequence-120715-        7557 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  VSVG reporter only (no hook) (RUSH system).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7557)
  AUTHORS   Boncompain G, Divoux S, Gareil N, de Forges H, Lescure A, Latreche 
            L, Mercanti V, Jollivet F, Raposo G, Perez F
  TITLE     Synchronization of secretory protein traffic in populations of 
            cells.
  JOURNAL   Nat Methods. 2012 Mar 11;9(5):493-8. doi: 10.1038/nmeth.1928.
  PUBMED    22406856
REFERENCE   2  (bases 1 to 7557)
  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/nmeth"; 
            journalName: "Nat Methods"; date: "2012-03-11- 11"; volume: "9"; 
            issue: "5"; pages: "493-8"
COMMENT     Sequence Label: VSVGwt-SBP-EGFP_puromycin
FEATURES             Location/Qualifiers
     source          1..7557
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(945..992,993..2477)
                     /codon_start=1
                     /product="vesicular stomatitis virus G glycoprotein"
                     /label=VSV-G
                     /note="Indiana strain"
                     /translation="MKCLLYLAFLFIGVNCKFTIVFPHNQKGNWKNVPSNYHYCPSSSD
                     LNWHNDLIGTALQVKMPKSHKAIQADGWMCHASKWVTTCDFRWYGPKYITHSIRSFTPS
                     VEQCKESIEQTKQGTWLNPGFPPQSCGYATVTDAEAVIVQVTPHHVLVDEYTGEWVDSQ
                     FINGKCSNYICPTVHNSTTWHSDYKVKGLCDSNLISMDITFFSEDGELSSLGKEGTGFR
                     SNYFAYETGDKACKMQYCKHWGVRLPSGVWFEMADKDLFAAARFPECPEGSSISAPSQT
                     SVDVSLIQDVERILDYSLCQETWSKIRAGLPISPVDLSYLAPKNPGTGPVFTIINGTLK
                     YFETRYIRVDIAAPILSRMVGMISGTTTERELWDDWAPYEDVEIGPNGVLRTSSGYKFP
                     LYMIGHGMLDSDLHLSSKAQVFEHPHIQDAASQLPDDESLFFGDTGLSKNPIELVEGWF
                     SSWKSSIASFFFIIGLIIGLFLVLRVGIHLCIKLKHTKKRQIYTDIEMNRLGK"
     CDS             2487..2600
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             join(2610..2612,2613..2615,2616..3326)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2610..2612,2613..2615,2616..3326)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2655..2676)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2916..2935)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3263..3284
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3436..3665
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3648..3667
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(3738..4312,4313..4315,4316..4324)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3905..3922)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4132..4151
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4344..4943
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4344..4363)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4840..4860
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5177..5298
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5214..5233)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5400..5416)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5400..5416)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5413..5435)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5424..5440
                     /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(5448..5454,5455..5472,5473..5478))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5493..5514
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5631..5648)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5802..6390)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5882..5901)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6561..7352,7353..7421))
                     /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     7184..7203
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7422..7526)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat cgatatctgc ggcctagcta gcgggaggcg cgccatgaag tgccttttgt
      961 acttagcctt tttattcatt ggggtgaatt gcaagttcac catagttttt ccacacaacc
     1021 aaaaaggaaa ctggaaaaat gttccttcta attaccatta ttgcccgtca agctcagatt
     1081 taaattggca taatgactta ataggcacag ccttacaagt caaaatgccc aagagtcaca
     1141 aggctattca agcagacggt tggatgtgtc atgcttccaa atgggtcact acttgtgatt
     1201 tccgctggta tggaccgaag tatataacac attccatccg atccttcact ccatctgtag
     1261 aacaatgcaa ggaaagcatt gaacaaacga aacaaggaac ttggctgaat ccaggcttcc
     1321 ctcctcaaag ttgtggatat gcaactgtga cggatgccga agcagtgatt gtccaggtga
     1381 ctcctcacca tgtgctggtt gatgaataca caggagaatg ggttgattca cagttcatca
     1441 acggaaaatg cagcaattac atatgcccca ctgtccataa ctctacaacc tggcattctg
     1501 actataaggt caaagggcta tgtgattcta acctcatttc catggacatc accttcttct
     1561 cagaggacgg agagctatca tccctaggaa aggagggcac agggttcaga agtaactact
     1621 ttgcttatga aactggagac aaggcctgca aaatgcagta ctgcaagcat tggggagtca
     1681 gactcccatc aggtgtctgg ttcgagatgg ctgataagga tctctttgct gcagccagat
     1741 tccctgaatg cccagaaggg tcaagtatct ctgctccatc tcagacctca gtggatgtaa
     1801 gtctcattca ggacgttgag aggatcttgg attattccct ctgccaagaa acctggagca
     1861 aaatcagagc gggtcttccc atctctccag tggatctcag ctatcttgct cctaaaaacc
     1921 caggaaccgg tcctgtcttt accataatca atggtaccct aaaatacttt gagaccagat
     1981 acatcagagt cgatattgct gctccaatcc tctcaagaat ggtcggaatg atcagtggaa
     2041 ctaccacaga aagggaactg tgggatgact gggcaccata tgaagacgtg gaaattggac
     2101 ccaatggagt tctgaggacc agttcaggat ataagtttcc tttatacatg attggacatg
     2161 gtatgttgga ctccgatctt catcttagct caaaggctca ggtgttcgaa catcctcaca
     2221 ttcaagacgc tgcttcgcaa cttcctgatg atgagagttt attttttggt gatactgggc
     2281 tatccaaaaa tccaatcgag cttgtagaag gttggttcag tagttggaaa agctctattg
     2341 cctctttttt ctttatcata gggttaatca ttggactatt cttggttctc cgagttggta
     2401 tccatctttg cattaaatta aagcacacca agaaaagaca gatttataca gacatagaga
     2461 tgaaccgact tggaaagctg aattccatgg acgagaagac cactggttgg cgaggtggac
     2521 acgttgttga aggactggct ggggaacttg aacaacttcg tgcacgactg gagcatcacc
     2581 cacaaggtca acgtgaacca cctgcaggta tggtgagcaa gggcgaggag ctgttcaccg
     2641 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
     2701 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
     2761 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
     2821 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
     2881 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
     2941 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
     3001 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
     3061 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
     3121 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
     3181 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
     3241 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
     3301 ctctcggcat ggacgagctg tacaagggcc ggccttaagg cctcgagggc caattaatta
     3361 aggatccgcg gccgcataga taactgatcc agtgtgctgg aattaattcg ctgtctgcga
     3421 gggccagctg ttggggtgag tactccctct caaaagcggg catgacttct gcgctaagat
     3481 tgtcagtttc caaaaacgag gaggatttga tattcacctg gcccgcggtg atgcctttga
     3541 gggtggccgc gtccatctgg tcagaaaaga caatcttttt gttgtcaagc ttgaggtgtg
     3601 gcaggcttga gatctggcca tacacttgag tgacaatgac atccactttg cctttctctc
     3661 cacaggtgtc cactcccagg tccaactgca ggtcgagcat gcatctaggg cggccaattc
     3721 cgcccctctc cccccccccc ctctccctcc ccccccccta acgttactgg ccgaagccgc
     3781 ttggaataag gccggtgtgc gtttgtctat atgtgatttt ccaccatatt gccgtctttt
     3841 ggcaatgtga gggcccggaa acctggccct gtcttcttga cgagcattcc taggggtctt
     3901 tcccctctcg ccaaaggaat gcaaggtctg ttgaatgtcg tgaaggaagc agttcctctg
     3961 gaagcttctt gaagacaaac aacgtctgta gcgacccttt gcaggcagcg gaacccccca
     4021 cctggcgaca ggtgcctctg cggccaaaag ccacgtgtat aagatacacc tgcaaaggcg
     4081 gcacaacccc agtgccacgt tgtgagttgg atagttgtgg aaagagtcaa atggctctcc
     4141 tcaagcgtat tcaacaaggg gctgaaggat gcccagaagg taccccattg tatgggatct
     4201 gatctggggc ctcggtgcac atgctttaca tgtgtttagt cgaggttaaa aaaacgtcta
     4261 ggccccccga accacgggga cgtggttttc ctttgaaaaa cacgatgata agcttgccac
     4321 aacccacaag gagacgacct tccatgaccg agtacaagcc cacggtgcgc ctcgccaccc
     4381 gcgacgacgt cccccgggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
     4441 cgcgccacac cgtcgacccg gaccgccaca tcgagcgggt caccgagctg caagaactct
     4501 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg
     4561 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     4621 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     4681 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg
     4741 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     4801 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc
     4861 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     4921 tgacccgcaa gcccggtgcc tgacgcccgc cccacgaccc gcagcgcccg accgaaagga
     4981 gcgcacgacc ccatggctcc gaccgaagcc acccggggcg gccccgccga ccccgcaccc
     5041 gcccccgagg cccaccgact ctagataact gatcataatc agccatacca catttgtaga
     5101 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     5161 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5221 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5281 actcatcaat gtatcttaac gcgtcgagtg cattctagtt gtggtttgtc caaactcatc
     5341 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     5401 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     5461 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     5521 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     5581 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     5641 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5701 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5761 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5821 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5881 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5941 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc
     6001 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6061 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6121 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6181 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6241 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6301 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6361 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6421 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6481 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     6541 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     6601 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6661 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6721 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6781 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6841 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6901 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6961 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     7021 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7081 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7141 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     7201 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7261 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7321 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7381 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7441 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7501 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
