LOCUS       Exported                7871 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  expresses V5-tagged TurboID in the yeast cytosol.
ACCESSION   .
VERSION     .
KEYWORDS    TurboID-V5_pRS415
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7871)
  AUTHORS   Branon TC, Bosch JA, Sanchez AD, Udeshi ND, Svinkina T, Carr SA, 
            Feldman JL, Perrimon N, Ting AY
  TITLE     Efficient proximity labeling in living cells and organisms with 
            TurboID.
  JOURNAL   Nat Biotechnol. 2018 Aug 20. pii: nbt.4201. doi: 10.1038/nbt.4201.
  PUBMED    30125270
REFERENCE   2  (bases 1 to 7871)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7871
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    97..118
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        133..163
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    171..187
                     /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)."
     primer_bind     176..198
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     195..211
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     195..211
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     230..250
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        232..250
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        441..882
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    441..558
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     853..876
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             954..1913
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             1920..1961
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     terminator      1974..2221
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1990..2008)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2239..2258)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2240..2258)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2268..2285)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2268..2284)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2277..2299)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2425..2880
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2512..2531)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2722..2743
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3181..3588
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3589..4683
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     primer_bind     complement(5177..5196)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5296..5318
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5356..5374)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5416..5919
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5956..6060
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6061..6921
                     /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     complement(6279..6298)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7092..7680
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7581..7600
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7834..7851
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
       61 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      121 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      181 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
      241 tcactaaagg gaacaaaagc tggagctccg caaagaagtt taataatcat attacatggc
      301 attaccacca tatacatatc catatctaat cttacttata tgttgtggaa atgtaaagag
      361 ccccattatc ttagcctaaa aaaaccttct ctttggaact ttcagtaata cgcttaactg
      421 ctcattgcta tattgaagta cggattagaa gccgccgagc gggtgacagc cctccgaagg
      481 aagactctcc tccgtgcgtc ctcgtcttca ccggtcgcgt tcctgaaacg cagatgtgcc
      541 tcgcgccgca ctgctccgaa caataaagat tctacaatac tagcttttat ggttatgaag
      601 aggaaaaatt ggcagtaacc tggccccaca aaccttcaaa tgaacgaatc aaattaacaa
      661 ccataggatg ataatgcgat tagtttttta gccttatttc tggggtaatt aatcagcgaa
      721 gcgatgattt ttgatctatt aacagatata taaatgcaaa aactgcataa ccactttaac
      781 taatactttc aacattttcg gtttgtatta cttcttattc aaatgtaata aaagtatcaa
      841 caaaaaattg ttaatatacc tctatacttt aacgtcaagg agaaaaaacc ccggatcgaa
      901 ttctacttca tacattttca attaagatgg ctcgggatcc accggtcgcc accatgaaag
      961 acaatactgt gcctctgaag ctgatcgctc tcctggctaa tggcgagttc catagtggcg
     1021 aacagctggg agaaaccctg ggcatgtcca gggccgctat caacaagcac attcagactc
     1081 tgcgcgactg gggcgtggac gtgttcaccg tgcccggaaa gggctactct ctgcccgagc
     1141 ctatcccgct gctgaacgct aaacagattc tgggacagct ggacggcggg agcgtggcag
     1201 tcctgcctgt ggtcgactcc accaatcagt acctgctgga tcgaatcggc gagctgaaga
     1261 gtggggatgc ttgcattgca gaatatcagc aggcagggag aggaagcaga gggaggaaat
     1321 ggttctctcc ttttggagct aacctgtacc tgagtatgtt ttggcgcctg aagcggggac
     1381 cagcagcaat cggcctgggc ccggtcatcg gaattgtcat ggcagaagcg ctgcgaaagc
     1441 tgggagcaga caaggtgcga gtcaaatggc ccaatgacct gtatctgcag gatagaaagc
     1501 tggcaggcat cctggtggag ctggccggaa taacaggcga tgctgcacag atcgtcattg
     1561 gcgccgggat taacgtggct atgaggcgcg tggaggaaag cgtggtcaat cagggctgga
     1621 tcacactgca ggaagcaggg attaacctgg acaggaatac tctggccgct acgctgatcc
     1681 gagagctgcg ggcagccctg gaactgttcg agcaggaagg cctggctcca tatctgccac
     1741 ggtgggagaa gctggataac ttcatcaata gacccgtgaa gctgatcatt ggggacaaag
     1801 agattttcgg gattagccgg gggattgata aacagggagc cctgctgctg gaacaggacg
     1861 gagttatcaa accctggatg ggcggagaaa tcagtctgcg gtctgccgaa aaggctagcg
     1921 gcaagcccat ccccaacccc ctgctgggcc tggacagcac ctaatagctc gagtcatgta
     1981 attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga
     2041 aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt
     2101 aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg
     2161 taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg
     2221 cggccggtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
     2281 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     2341 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     2401 ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     2461 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     2521 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     2581 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2641 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2701 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     2761 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     2821 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     2881 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatcgaccgg
     2941 tcgaggagaa cttctagtat atctacatac ctaatattat tgccttatta aaaatggaat
     3001 cccaacaatt acatcaaaat ccacattctc ttcaaaatca attgtcctgt acttccttgt
     3061 tcatgtgtgt tcaaaaacgt tatatttata ggataattat actctatttc tcaacaagta
     3121 attggttgtt tggccgagcg gtctaaggcg cctgattcaa gaaatatctt gaccgcagtt
     3181 aactgtggga atactcaggt atcgtaagat gcaagagttc gaatctctta gcaaccatta
     3241 tttttttcct caacataacg agaacacaca ggggcgctat cgcacagaat caaattcgat
     3301 gactggaaat tttttgttaa tttcagaggt cgcctgacgc atataccttt ttcaactgaa
     3361 aaattgggag aaaaaggaaa ggtgagagcg ccggaaccgg cttttcatat agaatagaga
     3421 agcgttcatg actaaatgct tgcatcacaa tacttgaagt tgacaatatt atttaaggac
     3481 ctattgtttt ttccaatagg tggttagcaa tcgtcttact ttctaacttt tcttaccttt
     3541 tacatttcag caatatatat atatatattt caaggatata ccattctaat gtctgcccct
     3601 aagaagatcg tcgttttgcc aggtgaccac gttggtcaag aaatcacagc cgaagccatt
     3661 aaggttctta aagctatttc tgatgttcgt tccaatgtca agttcgattt cgaaaatcat
     3721 ttaattggtg gtgctgctat cgatgctaca ggtgttccac ttccagatga ggcgctggaa
     3781 gcctccaaga aggctgatgc cgttttgtta ggtgctgtgg gtggtcctaa atggggtacc
     3841 ggtagtgtta gacctgaaca aggtttacta aaaatccgta aagaacttca attgtacgcc
     3901 aacttaagac catgtaactt tgcatccgac tctcttttag acttatctcc aatcaagcca
     3961 caatttgcta aaggtactga cttcgttgtt gtcagagaat tagtgggagg tatttacttt
     4021 ggtaagagaa aggaagacga tggtgatggt gtcgcttggg atagtgaaca atacaccgtt
     4081 ccagaagtgc aaagaatcac aagaatggcc gctttcatgg ccctacaaca tgagccacca
     4141 ttgcctattt ggtccttgga taaagctaat gttttggcct cttcaagatt atggagaaaa
     4201 actgtggagg aaaccatcaa gaacgaattc cctacattga aggttcaaca tcaattgatt
     4261 gattctgccg ccatgatcct agttaagaac ccaacccacc taaatggtat tataatcacc
     4321 agcaacatgt ttggtgatat catctccgat gaagcctccg ttatcccagg ttccttgggt
     4381 ttgttgccat ctgcgtcctt ggcctctttg ccagacaaga acaccgcatt tggtttgtac
     4441 gaaccatgcc acggttctgc tccagatttg ccaaagaata aggtcaaccc tatcgccact
     4501 atcttgtctg ctgcaatgat gttgaaattg tcattgaact tgcctgaaga aggtaaggcc
     4561 attgaagatg cagttaaaaa ggttttggat gcaggtatca gaactggtga tttaggtggt
     4621 tccaacagta ccaccgaagt cggtgatgct gtcgccgaag aagttaagaa aatccttgct
     4681 taaaaagatt ctcttttttt atgatatttg tacataaact ttataaatga aattcataat
     4741 agaaacgaca cgaaattaca aaatggaata tgttcatagg gtagacgaaa ctatatacgc
     4801 aatctacata catttatcaa gaaggagaaa aaggaggatg taaaggaata caggtaagca
     4861 aattgatact aatggctcaa cgtgataagg aaaaagaatt gcactttaac attaatattg
     4921 acaaggagga gggcaccaca caaaaagtta ggtgtaacag aaaatcatga aactatgatt
     4981 cctaatttat atattggagg attttctcta aaaaaaaaaa aatacaacaa ataaaaaaca
     5041 ctcaatgacc tgaccatttg atggagttta agtcaatacc ttcttgaacc atttcccata
     5101 atggtgaaag ttccctcaag aattttactc tgtcagaaac ggccttaacg acgtagtcga
     5161 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     5221 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     5281 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     5341 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     5401 tggtttctta ggacggatcg cttgcctgta acttacacgc gcctcgtatc ttttaatgat
     5461 ggaataattt gggaatttac tctgtgttta tttattttta tgttttgtat ttggatttta
     5521 gaaagtaaat aaagaaggta gaagagttac ggaatgaaga aaaaaaaata aacaaaggtt
     5581 taaaaaattt caacaaaaag cgtactttac atatatattt attagacaag aaaagcagat
     5641 taaatagata tacattcgat taacgataag taaaatgtaa aatcacagga ttttcgtgtg
     5701 tggtcttcta cacagacaag atgaaacaat tcggcattaa tacctgagag caggaagagc
     5761 aagataaaag gtagtatttg ttggcgatcc ccctagagtc ttttacatct tcggaaaaca
     5821 aaaactattt tttctttaat ttcttttttt actttctatt tttaatttat atatttatat
     5881 taaaaaattt aaattataat tatttttata gcacgtgatg aaaaggaccc aggtggcact
     5941 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     6001 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     6061 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     6121 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     6181 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     6241 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     6301 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     6361 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     6421 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     6481 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     6541 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     6601 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     6661 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     6721 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     6781 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     6841 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     6901 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     6961 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     7021 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     7081 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     7141 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     7201 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     7261 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     7321 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     7381 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     7441 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     7501 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     7561 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     7621 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     7681 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     7741 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     7801 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     7861 gagcgcccaa t
//
