LOCUS       Exported                5927 bp ds-DNA     circular SYN 14-SEP-2020
DEFINITION  Integrating vector for expressing the E2-Crimson version of ESCargo 
            in yeast cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5927)
  AUTHORS   Glick Lab
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Sep 17, 2020 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize with EcoRV to integrate at LEU2.
COMMENT     Alias: YIplac128-pOst1-APVNTT-E2-Crimson-FKBP-LV(C22V)
FEATURES             Location/Qualifiers
     source          1..5927
                     /organism="synthetic DNA construct"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
     promoter        1225..1711
                     /label=TPI1 promoter
                     /note="strong constitutive yeast promoter"
     CDS             1722..1787
                     /codon_start=1
                     /product="signal sequence of S. cerevisiae Ost1"
                     /label=pOst1
                     /note="drives cotranslational translocation"
                     /translation="MRQVWFSWIVGLFLCFFNVSSA"
     CDS             1788..1805
                     /codon_start=1
                     /product="tripeptide ER export signal followed by 
                     tripeptide N-glycosylation signal"
                     /label=APVNTT
                     /translation="APVNTT"
     CDS             1806..2477
                     /codon_start=1
                     /product="far-red noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2009)"
                     /label=E2-Crimson
                     /note="mammalian codon-optimized"
                     /translation="DSTENVIKPFMRFKVHMEGSVNGHEFEIEGVGEGKPYEGTQTAKL
                     QVTKGGPLPFAWDILSPQFFYGSKAYIKHPADIPDYLKQSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGTLIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERNYPRDGVLKGENHMA
                     LKLKGGGHYLCEFKSIYMAKKPVKLPGYHYVDYKLDITSHNEDYTVVEQYERAEARHHL
                     FQ"
     CDS             2490..2810
                     /codon_start=1
                     /product="cysteine-free improved reversibly dimerizing 
                     mutant of FK506-binding protein FKBP12 (Barrero et al., 
                     2016; Casler et al., 2019)"
                     /label=FKBP-LV(C22V)
                     /translation="GVQVETISPGDGRTFPKRGQTVVVHYTGMLEDGKKLDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGVIPPHATLVFDVELL
                     KLE"
     terminator      2824..3071
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(3491..4079)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number colE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4250..5110)
                     /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"
     promoter        complement(5111..5215)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5303..5707
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             join(5708..5927,1..875)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
ORIGIN
        1 taggtgctgt gggtggtcct aaatggggta caggtagtgt tagacctgaa caaggtttac
       61 taaaaatccg taaagaactt caattgtacg ccaacttaag accatgtaac tttgcatccg
      121 actctctttt agacttatct ccaatcaagc cacaatttgc taaaggtact gacttcgttg
      181 ttgtcagaga attagtggga ggtatttact ttggtaagag aaaggaagac gatggtgatg
      241 gtgtcgcttg ggatagtgaa caatacaccg ttccagaagt gcaaagaatc acaagaatgg
      301 ccgctttcat ggccctacaa catgagccac cattgcctat ttggtccttg gataaagcta
      361 atgttttggc ctcttcaaga ttatggagaa aaactgtgga ggaaaccatc aagaacgaat
      421 ttcctacatt gaaggttcaa catcaattga ttgattctgc cgccatgatc ctagttaaga
      481 acccaaccca cctaaatggt attataatca ccagcaacat gtttggtgat atcatctccg
      541 atgaagcctc cgttatccca ggttccttgg gtttgttgcc atctgcgtcc ttggcctctt
      601 tgccagacaa gaacaccgca tttggtttgt acgaaccatg ccacggttct gctccagatt
      661 tgccaaagaa taaggttgac cctatcgcca ctatcttgtc tgctgcaatg atgttgaaat
      721 tgtcattgaa cttgcctgaa gaaggtaagg ccattgaaga tgcagttaaa aaggttttgg
      781 atgcaggtat cagaactggt gatttaggtg gttccaacag taccaccgaa gtcggtgatg
      841 ctgtcgccga agaagttaag aaaatccttg cttaaaaaga ttctcttttt ttatgatatt
      901 tgtacataaa ctttataaat gaaattcata atagaaacga cacgaaatta caaaatggaa
      961 tatgttcata gggtaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct
     1021 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     1081 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
     1141 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     1201 taaggagaaa ataccgcatc aggcggacct taatacattc agacacttct gcggtatcac
     1261 cctacttatt cccttcgaga ttatatctag gaacccatca ggttggtgga agattacccg
     1321 ttctaagact tttcagcttc ctctattgat gttacacctg gacacccctt ttctggcatc
     1381 cagtttttaa tcttcagtgg catgtgagat tctccgaaat taattaaagc aatcacacaa
     1441 ttctctcgga taccacctcg gttgaaactg acaggtggtt tgttacgcat gctaatgcaa
     1501 aggagcctat atacctttgg ctcggctgct gtaacaggga atataaaggg cagcataatt
     1561 taggagttta gtgaacttgc aacatttact attttccctt cttacgtaaa tatttttctt
     1621 tttaattcta aatcaatctt tttcaatttt ttgtttgtat tcttttcttg cttaaatcta
     1681 taactacaaa aaacacatac ataaactaaa agaattcaaa aatgaggcag gtttggttct
     1741 cttggattgt gggattgttc ctatgttttt tcaacgtgtc ttctgctgcg ccggtgaaca
     1801 ccaccgatag cactgagaac gtcatcaagc ccttcatgcg cttcaaggtg cacatggagg
     1861 gctccgtgaa cggccacgag ttcgagatcg agggcgtggg cgagggcaag ccctacgagg
     1921 gcacccagac cgccaagctg caagtgacca agggcggccc cctgcccttc gcctgggaca
     1981 tcctgtcccc ccagttcttc tacggctcca aggcgtacat caagcacccc gccgacatcc
     2041 ccgactacct caagcagtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
     2101 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc accctcatct
     2161 accacgtgaa gttcatcggc gtgaacttcc cctccgacgg ccccgtaatg cagaagaaga
     2221 ctctgggctg ggagccctcc actgagcgca actacccccg cgacggcgtg ctgaagggcg
     2281 agaaccacat ggcgctgaag ctgaagggcg gcggccacta cctgtgtgag ttcaagtcca
     2341 tctacatggc caagaagccc gtgaagctgc ccggctacca ctacgtggac tacaagctcg
     2401 acatcacctc ccacaacgag gactacaccg tggtggagca gtacgagcgc gccgaggccc
     2461 gccaccacct gttccaggga tcctctagag gagtgcaggt ggaaaccatc tccccaggag
     2521 acgggcgcac cttccccaag cgcggccaga ccgttgtggt gcactacacc gggatgcttg
     2581 aagatggaaa gaaactagat tcctcccggg acagaaacaa gccctttaag tttatgctag
     2641 gcaagcagga ggtgatccga ggctgggaag aaggggttgc ccagatgagt gtgggtcaga
     2701 gagccaaact gactatatct ccagattatg cctatggtgc cactgggcac ccaggcgtta
     2761 tcccaccaca tgccactctc gtcttcgatg tggagcttct aaaactggaa taaactagag
     2821 tcatcatgta attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa
     2881 ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta
     2941 tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg
     3001 tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg
     3061 ctttaatttg caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3121 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3181 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3241 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3301 attgggcgtc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3361 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3421 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3481 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3541 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3601 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3661 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3721 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3781 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3841 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     3901 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     3961 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4021 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4081 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4141 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4201 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4261 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4321 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4381 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4441 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4501 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4561 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4621 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     4681 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4741 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4801 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     4861 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     4921 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     4981 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5041 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5101 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5161 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5221 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     5281 aaaataggcg tatcacgagg ccaactgtgg gaatactcag gtatcgtaag atgcaagagt
     5341 tcgaatctct tagcaaccat tatttttttc ctcaacataa cgagaacaca caggggcgct
     5401 atcgcacaga atcaaattcg atgactggaa attttttgtt aatttcagag gtcgcctgac
     5461 gcatatacct ttttcaactg aaaaattggg agaaaaagga aaggtgagag gccggaaccg
     5521 gcttttcata tagaatagag aagcgttcat gactaaatgc ttgcatcaca atacttgaag
     5581 ttgacaatat tatttaagga cctattgttt tttccaatag gtggttagca atcgtcttac
     5641 tttctaactt ttcttacctt ttacatttca gcaatatata tatatatttc aaggatatac
     5701 cattctaatg tctgccccta agaagatcgt cgttttgcca ggtgaccacg ttggtcaaga
     5761 aatcacagcc gaagccatta aggttcttaa agctatttct gatgttcgtt ccaatgtcaa
     5821 gttcgatttc gaaaatcatt taattggtgg tgctgctatc gatgctacag gtgtcccact
     5881 tccagatgag gcgctggaag cctccaagaa ggttgatgcc gttttgt
//
