LOCUS       sequence-492557-        8033 bp    DNA     circular SYN 11-NOV-2025
DEFINITION  synchronize trafficking of VSVG from the ER (RUSH system).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8033)
  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 8033)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 11, 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: Str-Ii_VSVG-SBP-EGFP
FEATURES             Location/Qualifiers
     source          1..8033
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(48..67)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        239..618
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..822
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     773..793
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     867..886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        867..885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1459..1485
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          2220..2449
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2432..2451
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(2522..3096,3097..3099,3100..3108)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2689..2706)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2916..2935
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3122..3169,3170..4654)
                     /codon_start=1
                     /product="vesicular stomatitis virus G glycoprotein"
                     /label=VSV-G
                     /note="Indiana strain"
                     /translation="MKCLLYLAFLFIGVNCKFTIVFPHNQKGNWKNVPSNYHYCPSSSD
                     LNWHNDLIGTALQVKMPKSHKAIQADGWMCHASKWVTTCDFRWYGPKYITHSIRSFTPS
                     VEQCKESIEQTKQGTWLNPGFPPQSCGYATVTDAEAVIVQVTPHHVLVDEYTGEWVDSQ
                     FINGKCSNYICPTVHNSTTWHSDYKVKGLCDSNLISMDITFFSEDGELSSLGKEGTGFR
                     SNYFAYETGGKACKMQYCKHWGVRLPSGVWFEMADKDLFAAARFPECPEGSSISAPSQT
                     SVDVSLIQDVERILDYSLCQETWSKIRAGLPISPVDLSYLAPKNPGTGPAFTIINGTLK
                     YFETRYIRVDIAAPILSRMVGMISGTTTERELWDDWAPYEDVEIGPNGVLRTSSGYKFP
                     LYMIGHGMLDSDLHLSSKAQVFEHPHIQDAASQLPDDESLFFGDTGLSKNPIELVEGWF
                     SSWKSSIASFFFIIGLIIGLFLVLRVGIHLCIKLKHTKKRQIYTDIEMNRLGK"
     CDS             4664..4777
                     /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(4787..4789,4790..4792,4793..5503)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4787..4789,4790..4792,4793..5503)
                     /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(4832..4853)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5093..5112)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5440..5461
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5657..5778
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5694..5713)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5880..5896)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5880..5896)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5893..5915)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5904..5920
                     /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(5928..5934,5935..5952,5953..5958))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5973..5994
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6111..6128)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6282..6870)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6362..6381)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7041..7832,7833..7901))
                     /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     7664..7683
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7902..8006)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
       61 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
      121 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
      181 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
      241 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      301 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      361 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      421 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      481 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      541 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      601 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      661 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      721 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      781 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      841 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagcttgg
      901 taccgagctc ggatcgatat catgcaccgg aggagatcac gctcttgtag ggaggaccag
      961 aaacctgtca ccggtgaccc tagcaaagac tcaaaagctc aggtgtccgc tgccgaggct
     1021 ggcattactg gaacatggta caatcagctc gggagcacct ttattgtgac tgctggagcc
     1081 gatggagccc tcaccggaac atacgaatct gctgtgggaa acgccgaatc acggtacgtc
     1141 ctcactggcc gatacgatag tgcccctgcc accgacggat ctgggactgc cctgggatgg
     1201 actgtcgctt ggaaaaacaa ctaccggaat gctcattctg ccacaacatg gagtggacag
     1261 tacgtgggag gcgctgaggc tagaatcaat acacagtggc tgctcacatc tggcacaacc
     1321 gaggcaaatg cttggaaatc caccctggtg ggacatgaca cattcaccaa agtgaaaccc
     1381 tccgccgctt caatcgatgc cgccaaaaaa gccggagtca acaacggcaa tcctctggat
     1441 gccgtccagc aggtcgacta tccgtacgac gtaccagact acgcagtcgg accgatggac
     1501 gatcagaggg acctcattag caacaacgaa cagctgccta tgctgggacg gcgacctgga
     1561 gcccctgaat ccaaatgctc taggggagca ctgtacactg gcttctccat tctcgtgaca
     1621 ctgctgctgg ccgggcaggc tactactgct tacttcctgt accagcagca ggggcggctg
     1681 gacaaactca ctgtgacatc tcagaacctc cagctggaaa atctgaggat gaaactgccc
     1741 aaacccccta aacccgtgtc caaaatgagg atggccacac ctctgctcat gcaggcactg
     1801 ccaatgggag ccctgcccca ggggcccatg cagaatgcca ccaagtatgg caacatgaca
     1861 gaggaccatg tgatgcacct gctccagaat gctgaccccc tgaaggtgta cccgccactg
     1921 aaggggagct tcccggagaa cctgagacac cttaagaaca ccatggagac catagactgg
     1981 aaggtctttg agagctggat gcaccattgg ctcctgtttg aaatgagcag gcactccttg
     2041 gagcaaaagc ccactgacgc tccaccgaaa gagtcactgg aactggagga cccgtcttct
     2101 gggctgggtg tgaccaagca ggatctgggc ccagtcccca tgtgaggatc cgcggccgca
     2161 tagataactg atccagtgtg ctggaattaa ttcgctgtct gcgagggcca gctgttgggg
     2221 tgagtactcc ctctcaaaag cgggcatgac ttctgcgcta agattgtcag tttccaaaaa
     2281 cgaggaggat ttgatattca cctggcccgc ggtgatgcct ttgagggtgg ccgcgtccat
     2341 ctggtcagaa aagacaatct ttttgttgtc aagcttgagg tgtggcaggc ttgagatctg
     2401 gccatacact tgagtgacaa tgacatccac tttgcctttc tctccacagg tgtccactcc
     2461 caggtccaac tgcaggtcga gcatgcatct agggcggcca attccgcccc tctccccccc
     2521 ccccctctcc ctcccccccc cctaacgtta ctggccgaag ccgcttggaa taaggccggt
     2581 gtgcgtttgt ctatatgtta ttttccacca tattgccgtc ttttggcaat gtgagggccc
     2641 ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg tctttcccct ctcgccaaag
     2701 gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc tctggaagct tcttgaagac
     2761 aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc cccacctggc gacaggtgcc
     2821 tctgcggcca aaagccacgt gtataagata cacctgcaaa ggcggcacaa ccccagtgcc
     2881 acgttgtgag ttggatagtt gtggaaagag tcaaatggct ctcctcaagc gtattcaaca
     2941 aggggctgaa ggatgcccag aaggtacccc attgtatggg atctgatctg gggcctcggt
     3001 gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg tctaggcccc ccgaaccacg
     3061 gggacgtggt tttcctttga aaaacacgat gataagcttg ccacaacccg ggaggcgcgc
     3121 catgaagtgc cttttgtact tagccttttt attcattggg gtgaattgca agttcaccat
     3181 agtttttcca cacaaccaaa aaggaaactg gaaaaatgtt ccttctaatt accattattg
     3241 cccgtcaagc tcagatttaa attggcataa tgacttaata ggcacagcct tacaagtcaa
     3301 aatgcccaag agtcacaagg ctattcaagc agacggttgg atgtgtcatg cttccaaatg
     3361 ggtcactact tgtgatttcc gctggtatgg accgaagtat ataacacatt ccatccgatc
     3421 cttcactcca tctgtagaac aatgcaagga aagcattgaa caaacgaaac aaggaacttg
     3481 gctgaatcca ggcttccctc ctcaaagttg tggatatgca actgtgacgg atgccgaagc
     3541 agtgattgtc caggtgactc ctcaccatgt gctggttgat gaatacacag gagaatgggt
     3601 tgattcacag ttcatcaacg gaaaatgcag caattacata tgccccactg tccataactc
     3661 tacaacctgg cattctgact ataaggtcaa agggctatgt gattctaacc tcatttccat
     3721 ggacatcacc ttcttctcag aggacggaga gctatcatcc ctgggaaagg agggcacagg
     3781 gttcagaagt aactactttg cttatgaaac tggaggcaag gcctgcaaaa tgcaatactg
     3841 caagcattgg ggagtcagac tcccatcagg tgtctggttc gagatggctg ataaggatct
     3901 ctttgctgca gccagattcc ctgaatgccc agaagggtca agtatctctg ctccatctca
     3961 gacctcagtg gatgtaagtc taattcagga cgttgagagg atcttggatt attccctctg
     4021 ccaagaaacc tggagcaaaa tcagagcggg tcttccaatc tctccagtgg atctcagcta
     4081 tcttgctcct aaaaacccag gaaccggtcc tgctttcacc ataatcaatg gtaccctaaa
     4141 atactttgag accagataca tcagagtcga tattgctgct ccaatcctct caagaatggt
     4201 cggaatgatc agtggaacta ccacagaaag ggaactgtgg gatgactggg caccatatga
     4261 agacgtggaa attggaccca atggagttct gaggaccagt tcaggatata agtttccttt
     4321 atacatgatt ggacatggta tgttggactc cgatcttcat cttagctcaa aggctcaggt
     4381 gttcgaacat cctcacattc aagacgctgc ttcgcaactt cctgatgatg agagtttatt
     4441 ttttggtgat actgggctat ccaaaaatcc aatcgagctt gtagaaggtt ggttcagtag
     4501 ttggaaaagc tctattgcct cttttttctt tatcataggg ttaatcattg gactattctt
     4561 ggttctccga gttggtatcc atctttgcat taaattaaag cacaccaaga aaagacagat
     4621 ttatacagac atagagatga accgacttgg aaagctgaat tccatggacg agaagaccac
     4681 tggttggcga ggtggacacg ttgttgaagg actggctggg gaacttgaac aacttcgtgc
     4741 acgactggag catcacccac aaggtcaacg tgaaccacct gcaggtatgg tgagcaaagg
     4801 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     4861 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     4921 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     4981 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
     5041 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     5101 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     5161 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     5221 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     5281 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     5341 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     5401 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     5461 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagggccggc cttaaggcct
     5521 cgagggccaa ttaattaact ctagataact gatcataatc agccatacca catttgtaga
     5581 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     5641 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5701 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5761 actcatcaat gtatcttaac gcgtcgagtg cattctagtt gtggtttgtc caaactcatc
     5821 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     5881 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     5941 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     6001 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     6061 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     6121 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     6181 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     6241 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     6301 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     6361 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     6421 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     6481 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6541 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6601 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6661 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6721 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6781 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6841 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     6901 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     6961 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     7021 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     7081 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     7141 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     7201 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     7261 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     7321 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     7381 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     7441 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     7501 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     7561 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     7621 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     7681 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     7741 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     7801 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     7861 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     7921 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     7981 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tga
//
