LOCUS       Exported                5183 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 5183)
  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 TRP1.
COMMENT     Alias: YIplac204-pOst1-APVNTT-E2-Crimson-FKBP-LV(C22V)
FEATURES             Location/Qualifiers
     source          1..5183
                     /organism="synthetic DNA construct"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
     rep_origin      complement(183..771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number colE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(942..1802)
                     /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(1803..1907)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1995..2096
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             2097..2771
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        3099..3585
                     /label=TPI1 promoter
                     /note="strong constitutive yeast promoter"
     CDS             3596..3661
                     /codon_start=1
                     /product="signal sequence of S. cerevisiae Ost1"
                     /label=pOst1
                     /note="drives cotranslational translocation"
                     /translation="MRQVWFSWIVGLFLCFFNVSSA"
     CDS             3662..3679
                     /codon_start=1
                     /product="tripeptide ER export signal followed by 
                     tripeptide N-glycosylation signal"
                     /label=APVNTT
                     /translation="APVNTT"
     CDS             3680..4351
                     /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             4364..4684
                     /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      4698..4945
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
ORIGIN
        1 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
       61 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      121 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
      181 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
      241 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
      301 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
      361 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
      421 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
      481 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
      541 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
      601 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
      661 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
      721 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
      781 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
      841 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
      901 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
      961 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1021 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1081 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1141 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     1201 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     1261 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     1321 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     1381 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     1441 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     1501 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     1561 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     1621 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     1681 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     1741 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     1801 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     1861 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     1921 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     1981 cgtatcacga ggccaattcg gtcgaaaaaa gaaaaggaga gggccaagag ggagggcatt
     2041 ggtgactatt gagcacgtga gtatacgtga ttaagcacac aaaggcagct tggagtatgt
     2101 ctgttattaa tttcacaggt agttctggtc cattggtgaa agtttgcggc ttgcagagca
     2161 cagaggccgc agaatgtgca ctagattccg atgctgactt gctgggtatt atatgtgtgc
     2221 ccaatagaaa gagaacaatt gacccggtta ttgcaaggaa aatttcaagt cttgtaaaag
     2281 catataaaaa tagttcaggc actccgaaat acttggttgg cgtgtttcgt aatcaaccta
     2341 aggaggatgt tttggctctg gtcaatgatt acggcattga tatcgtccaa ctgcatggag
     2401 atgagtcgtg gcaagaatac caagagttcc tcggtttgcc agttattaaa agactcgtat
     2461 ttccaaaaga ctgcaacata ctactcagtg cagcttcaca gaaacctcat tcgtttattc
     2521 ccttgtttga ttcagaagca ggtgggacag gtgaactttt ggattggaac tcgatttctg
     2581 actgggttgg aaggcaagag agccccgaga gcttacattt tatgttagct ggtggactga
     2641 cgccagaaaa tgttggtgat gcgcttagat taaatggcgt tattggtgtt gatgtaagcg
     2701 gaggtgtgga gacaaatggt gtaaaagact ctaacaaaat agcaaatttc gtcaaaaatg
     2761 ctaagaaata ggttattact gagtagtatt tatttaagta ttgtttgtgc acttgcctgc
     2821 aagccttttg aaaagcaagc ataaaagatc gccctttcgt ctcgcgcgtt tcggtgatga
     2881 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     2941 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     3001 gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat
     3061 accgcacaga tgcgtaagga gaaaataccg catcaggcgg accttaatac attcagacac
     3121 ttctgcggta tcaccctact tattcccttc gagattatat ctaggaaccc atcaggttgg
     3181 tggaagatta cccgttctaa gacttttcag cttcctctat tgatgttaca cctggacacc
     3241 ccttttctgg catccagttt ttaatcttca gtggcatgtg agattctccg aaattaatta
     3301 aagcaatcac acaattctct cggataccac ctcggttgaa actgacaggt ggtttgttac
     3361 gcatgctaat gcaaaggagc ctatatacct ttggctcggc tgctgtaaca gggaatataa
     3421 agggcagcat aatttaggag tttagtgaac ttgcaacatt tactattttc ccttcttacg
     3481 taaatatttt tctttttaat tctaaatcaa tctttttcaa ttttttgttt gtattctttt
     3541 cttgcttaaa tctataacta caaaaaacac atacataaac taaaagaatt caaaaatgag
     3601 gcaggtttgg ttctcttgga ttgtgggatt gttcctatgt tttttcaacg tgtcttctgc
     3661 tgcgccggtg aacaccaccg atagcactga gaacgtcatc aagcccttca tgcgcttcaa
     3721 ggtgcacatg gagggctccg tgaacggcca cgagttcgag atcgagggcg tgggcgaggg
     3781 caagccctac gagggcaccc agaccgccaa gctgcaagtg accaagggcg gccccctgcc
     3841 cttcgcctgg gacatcctgt ccccccagtt cttctacggc tccaaggcgt acatcaagca
     3901 ccccgccgac atccccgact acctcaagca gtccttcccc gagggcttca agtgggagcg
     3961 cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct ccctgcagga
     4021 cggcaccctc atctaccacg tgaagttcat cggcgtgaac ttcccctccg acggccccgt
     4081 aatgcagaag aagactctgg gctgggagcc ctccactgag cgcaactacc cccgcgacgg
     4141 cgtgctgaag ggcgagaacc acatggcgct gaagctgaag ggcggcggcc actacctgtg
     4201 tgagttcaag tccatctaca tggccaagaa gcccgtgaag ctgcccggct accactacgt
     4261 ggactacaag ctcgacatca cctcccacaa cgaggactac accgtggtgg agcagtacga
     4321 gcgcgccgag gcccgccacc acctgttcca gggatcctct agaggagtgc aggtggaaac
     4381 catctcccca ggagacgggc gcaccttccc caagcgcggc cagaccgttg tggtgcacta
     4441 caccgggatg cttgaagatg gaaagaaact agattcctcc cgggacagaa acaagccctt
     4501 taagtttatg ctaggcaagc aggaggtgat ccgaggctgg gaagaagggg ttgcccagat
     4561 gagtgtgggt cagagagcca aactgactat atctccagat tatgcctatg gtgccactgg
     4621 gcacccaggc gttatcccac cacatgccac tctcgtcttc gatgtggagc ttctaaaact
     4681 ggaataaact agagtcatca tgtaattagt tatgtcacgc ttacattcac gccctccccc
     4741 cacatccgct ctaaccgaaa aggaaggagt tagacaacct gaagtctagg tccctattta
     4801 tttttttata gttatgttag tattaagaac gttatttata tttcaaattt ttcttttttt
     4861 tctgtacaga cgcgtgtacg catgtaacat tatactgaaa accttgcttg agaaggtttt
     4921 gggacgctcg aaggctttaa tttgcaagct tggcgtaatc atggtcatag ctgtttcctg
     4981 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     5041 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     5101 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     5161 gaggcggttt gcgtattggg cgc
//
