LOCUS       Exported                7065 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  to synchronize transport of secretory soluble GFP from the ER (RUSH 
            system).
ACCESSION   .
VERSION     .
KEYWORDS    Ii-Str_ssSBP-EGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7065)
  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 7065)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7065
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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             1401..1427
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          1946..2175
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2158..2177
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2233..2806
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2400..2417)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2627..2646
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     sig_peptide     2833..2892
                     /product="human interleukin-2 signal sequence"
                     /label=IL-2 signal sequence
     CDS             2893..3003
                     /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="DEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             3013..3729
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3013..3729
                     /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(3058..3079)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3319..3338)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3666..3687
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3853..3974
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3890..3909)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3944..3963
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3981..4436)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4118..4139)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4330..4349
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4463..4567
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4569..4926
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(4592..4612)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4777..4912
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4839..4858
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4961..5755
                     /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(5015..5034)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5625..5644
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5943..5962)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5987..6034
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6363..6951
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6852..6871
                     /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 ccggtcgtac gtctccggat tcgaattaat tcggatccgt gatgcacagg aggagagcca
      661 gggcctgtcg ggaagatcaa aagccagtca tcgatgatca gcgcgacctt atctccaaca
      721 atgagcaact gcccatgctg ggccggcgcc ctggggcccc ggagagcaag tgcagccgcg
      781 gagccctgta cacaggcttt tccatcctgg tgactctgct cctcgctggc caggccacca
      841 ccgcctactt cctgtaccag cagcagggcc ggctggacaa actgacagtc acctcccaga
      901 acctgcagct cgagatggac cctagcaaag actcaaaagc tcaggtgtcc gctgccgagg
      961 ctggcattac tggaacatgg tacaatcagc tcgggagcac ctttattgtg actgctggag
     1021 ccgatggagc cctcaccgga acatacgaat ctgctgtggg aaacgccgaa tcacggtacg
     1081 tcctcactgg ccgatacgat agtgcccctg ccaccgacgg atctgggact gccctgggat
     1141 ggactgtcgc ttggaaaaac aactaccgga atgctcattc tgccacaaca tggagtggac
     1201 agtacgtggg aggcgctgag gctagaatca atacacagtg gctgctcaca tctggcacaa
     1261 ccgaggcaaa tgcttggaaa tccaccctgg tgggacatga cacattcacc aaagtgaaac
     1321 cctccgccgc ttcaatcgat gccgccaaaa aagccggagt caacaacggc aatcctctgg
     1381 atgccgtcca gcaggtcgac tatccgtacg acgtaccaga ctacgcactc gacctgcagc
     1441 tggagaacct gcgcatgaag cttcccaagc ctcccaagcc tgtgagcaag atgcgcatgg
     1501 ccaccccgct gctgatgcat gcgctgccca tgggagccct gccccagggg cccatgcaga
     1561 atgccaccaa gtatggcaac atgacagagg accatgtgat gcacctgctc cagaatgctg
     1621 accccctgaa ggtgtacccg ccactgaagg ggagcttccc ggagaacctg agacacctta
     1681 agaacaccat ggagaccata gactggaagg tctttgagag ctggatgcac cattggctcc
     1741 tgtttgaaat gagcaggcac tccttggagc aaaagcccac tgacgctcca ccgaaagagt
     1801 cactggaact ggaggacccg tcttctgggc tgggtgtgac caagcaggat ctgggcccag
     1861 tccccatgtg aggatccgcg gccgcataga taactgatcc agtgtgctgg aattaattcg
     1921 ctgtctgcga gggccagctg ttggggtgag tactccctct caaaagcggg catgacttct
     1981 gcgctaagat tgtcagtttc caaaaacgag gaggatttga tattcacctg gcccgcggtg
     2041 atgcctttga gggtggccgc gtccatctgg tcagaaaaga caatcttttt gttgtcaagc
     2101 ttgaggtgtg gcaggcttga gatctggcca tacacttgag tgacaatgac atccactttg
     2161 cctttctctc cacaggtgtc cactcccagg tccaactgca ggtcgagcat gcatctaggg
     2221 cggccaattc cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga
     2281 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa
     2341 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc
     2401 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc
     2461 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg
     2521 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca
     2581 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag
     2641 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct
     2701 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc
     2761 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaaccc
     2821 gggaggcgcg ccatgtacag gatgcaactc ctgtcttgca ttgcactaag tcttgcactt
     2881 gtcacgaatt ccgacgagaa gaccactggt tggcgaggtg gacacgttgt tgaaggactg
     2941 gctggggaac ttgaacaact tcgtgcacga ctggagcatc acccacaagg tcaacgtgaa
     3001 ccacctgcag gtatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     3061 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     3121 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     3181 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     3241 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     3301 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     3361 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     3421 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     3481 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     3541 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     3601 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     3661 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     3721 ctgtacaagg gtaactctag ataactgatc ataatcagcc ataccacatt tgtagaggtt
     3781 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     3841 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     3901 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     3961 atcaatgtat cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa
     4021 tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa
     4081 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact
     4141 attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc
     4201 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     4261 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     4321 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     4381 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg
     4441 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4501 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     4561 gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag
     4621 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4681 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     4741 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     4801 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     4861 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     4921 tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     4981 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     5041 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     5101 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct
     5161 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     5221 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     5281 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     5341 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     5401 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     5461 agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac
     5521 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     5581 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     5641 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     5701 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     5761 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     5821 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     5881 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta
     5941 actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga
     6001 cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc
     6061 tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc
     6121 ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg
     6181 gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt
     6241 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     6301 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     6361 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     6421 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     6481 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     6541 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     6601 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     6661 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     6721 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     6781 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     6841 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     6901 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     6961 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     7021 gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcat
//
