LOCUS       Exported                6482 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Luminal ER hook only (RUSH system).
ACCESSION   .
VERSION     .
KEYWORDS    Ii-Str_neomycin
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6482)
  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 6482)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6482
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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             1753..1779
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          2298..2527
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2510..2529
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    2585..3158
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2752..2769)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2979..2998
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3185..3979
                     /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(3239..3258)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3849..3868
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4106..4227
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4143..4162)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4329..4345)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4329..4345)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4342..4364)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4353..4369
                     /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(4377..4407)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4422..4443
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4560..4577)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4731..5319)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4811..4830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5490..6350)
                     /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     6113..6132
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6351..6455)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgtcgacgga tcgggagatc tcccgatccc ctatggtcga 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 ggactattta 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 ctgcggccta gctagcgctt aaggcctgtt aaccggtcgt
      961 acgtctccgg attcgaatta attcggatcc gtgatgcaca ggaggagagc cagggcctgt
     1021 cgggaagatc aaaagccagt catcgatgat cagcgcgacc ttatctccaa caatgagcaa
     1081 ctgcccatgc tgggccggcg ccctggggcc ccggagagca agtgcagccg cggagccctg
     1141 tacacaggct tttccatcct ggtgactctg ctcctcgctg gccaggccac caccgcctac
     1201 ttcctgtacc agcagcaggg ccggctggac aaactgacag tcacctccca gaacctgcag
     1261 ctcgagatgg accctagcaa agactcaaaa gctcaggtgt ccgctgccga ggctggcatt
     1321 actggaacat ggtacaatca gctcgggagc acctttattg tgactgctgg agccgatgga
     1381 gccctcaccg gaacatacga atctgctgtg ggaaacgccg aatcacggta cgtcctcact
     1441 ggccgatacg atagtgcccc tgccaccgac ggatctggga ctgccctggg atggactgtc
     1501 gcttggaaaa acaactaccg gaatgctcat tctgccacaa catggagtgg acagtacgtg
     1561 ggaggcgctg aggctagaat caatacacag tggctgctca catctggcac aaccgaggca
     1621 aatgcttgga aatccaccct ggtgggacat gacacattca ccaaagtgaa accctccgcc
     1681 gcttcaatcg atgccgccaa aaaagccgga gtcaacaacg gcaatcctct ggatgccgtc
     1741 cagcaggtcg actatccgta cgacgtacca gactacgcac tcgacctgca gctggagaac
     1801 ctgcgcatga agcttcccaa gcctcccaag cctgtgagca agatgcgcat ggccaccccg
     1861 ctgctgatgc aggcgctgcc catgggagcc ctgccccagg ggcccatgca gaatgccacc
     1921 aagtatggca acatgacaga ggaccatgtg atgcacctgc tccagaatgc tgaccccctg
     1981 aaggtgtacc cgccactgaa ggggagcttc ccggagaacc tgagacacct taagaacacc
     2041 atggagacca tagactggaa ggtctttgag agctggatgc accattggct cctgtttgaa
     2101 atgagcaggc actccttgga gcaaaagccc actgacgctc caccgaaaga gtcactggaa
     2161 ctggaggacc cgtcttctgg gctgggtgtg accaagcagg atctgggccc agtccccatg
     2221 tgaggatccg cggccgcata gataactgat ccagtgtgct ggaattaatt cgctgtctgc
     2281 gagggccagc tgttggggtg agtactccct ctcaaaagcg ggcatgactt ctgcgctaag
     2341 attgtcagtt tccaaaaacg aggaggattt gatattcacc tggcccgcgg tgatgccttt
     2401 gagggtggcc gcgtccatct ggtcagaaaa gacaatcttt ttgttgtcaa gcttgaggtg
     2461 tggcaggctt gagatctggc catacacttg agtgacaatg acatccactt tgcctttctc
     2521 tccacaggtg tccactccca ggtccaactg caggtcgagc atgcatctag ggcggccaat
     2581 tccgcccctc tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc
     2641 ggtgtgcgtt tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg
     2701 cccggaaacc tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca
     2761 aaggaatgca aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa
     2821 gacaaacaac gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt
     2881 gcctctgcgg ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt
     2941 gccacgttgt gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca
     3001 acaaggggct gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc
     3061 ggtgcacatg ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc
     3121 acggggacgt ggttttcctt tgaaaaacac gatgataagc ttgccacaac ccgggaggcg
     3181 cgccatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3241 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3301 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3361 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3421 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3481 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3541 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3601 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3661 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     3721 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3781 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     3841 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     3901 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     3961 tcttgacgag ttcttctgat taattaactc tagataactg atcataatca gccataccac
     4021 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     4081 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     4141 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4201 tttgtccaaa ctcatcaatg tatcttaacg cgtcgagtgc attctagttg tggtttgtcc
     4261 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     4321 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4381 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4441 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4501 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4561 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4621 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4681 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4741 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4801 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4861 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4921 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4981 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5041 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5101 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5161 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5221 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5281 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5341 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5401 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5461 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5521 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5581 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5641 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5701 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5761 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5821 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5881 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5941 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6001 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6061 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6121 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6181 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6241 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6301 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6361 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6421 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6481 ga
//
