LOCUS       sequence-472644-        7880 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ScVPS34 delta CBR1 L31-D55-3xFLAG undeer its own 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7880)
  AUTHORS   Rostislavleva K, Soler N, Ohashi Y, Zhang L, Pardon E, Burke JE, 
            Masson GR, Johnson C, Steyaert J, Ktistakis NT, Williams RL
  TITLE     Structure and flexibility of the endosomal Vps34 complex reveals the
            basis of its function on membranes.
  JOURNAL   Science. 2015 Oct 9;350(6257):aac7365. doi: 10.1126/science.aac7365.
  PUBMED    26450213
REFERENCE   2  (bases 1 to 7880)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science."; 
            date: "2015-10-9"; volume: "350(6257):aac7365. doi"; pages: " 
            10.1126/science.aac7365"
COMMENT     Sequence Label: pYO934
FEATURES             Location/Qualifiers
     source          1..7880
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        194..409
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             410..1213
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(1344..1799)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1481..1502)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1693..1712
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1925..1947
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1939..1956
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1940..1956
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1966..1985
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1966..1984
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2187..2198
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4769..4834
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     terminator      4851..5038
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4906..4930)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(5107..5127)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5107..5125)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5146..5162)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5146..5162)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5159..5181)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5170..5186
                     /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(join(5194..5200,5201..5218,5219..5224))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5239..5260
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5377..5394)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5548..6136)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5628..5647)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6307..7098,7099..7167))
                     /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     6930..6949
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7168..7272)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    7309..7812
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     7853..7871
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataccac cttttcaatt catcattttt tttttattct tttttttgat ttcggtttcc
      241 ttgaaatttt tttgattcgg taatctccga acagaaggaa gaacgaagga aggagcacag
      301 acttagattg gtatatatac gcatatgtag tgttgaagaa acatgaaatt gcccagtatt
      361 cttaacccaa ctgcacagaa caaaaacctg caggaaacga agataaatca tgtcgaaagc
      421 tacatataag gaacgtgctg ctactcatcc tagtcctgtt gctgccaagc tatttaatat
      481 catgcacgaa aagcaaacaa acttgtgtgc ttcattggat gttcgtacca ccaaggaatt
      541 actggagtta gttgaagcat taggtcccaa aatttgttta ctaaaaacac atgtggatat
      601 cttgactgat ttttccatgg agggcacagt taagccgcta aaggcattat ccgccaagta
      661 caatttttta ctcttcgaag acagaaaatt tgctgacatt ggtaatacag tcaaattgca
      721 gtactctgcg ggtgtataca gaatagcaga atgggcagac attacgaatg cacacggtgt
      781 ggtgggccca ggtattgtta gcggtttgaa gcaggcggca gaagaagtaa caaaggaacc
      841 tagaggcctt ttgatgttag cagaattgtc atgcaagggc tccctatcta ctggagaata
      901 tactaagggt actgttgaca ttgcgaagag cgacaaagat tttgttatcg gctttattgc
      961 tcaaagagac atgggtggaa gagatgaagg ttacgattgg ttgattatga cacccggtgt
     1021 gggtttagat gacaagggag acgcattggg tcaacagtat agaaccgtgg atgatgtggt
     1081 ctctacagga tctgacatta ttattgttgg aagaggacta tttgcaaagg gaagggatgc
     1141 taaggtagag ggtgaacgtt acagaaaagc aggctgggaa gcatatttga gaagatgcgg
     1201 ccagcaaaac taaaaaactg tattataagt aaatgcatgt atactaaact cacaaattag
     1261 agcttcaatt taattatatc agttattacc ctatgcggtg tgaaataccg cacagatgcg
     1321 taaggagaaa ataccgcatc aggaaattgt aagcgttaat attttgttaa aattcgcgtt
     1381 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     1441 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     1501 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     1561 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     1621 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     1681 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     1741 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     1801 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     1861 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     1921 ttttcccagt cacgacgttg taaaacgacg gccagtgagc gcgcgtaata cgactcacta
     1981 tagggcgaat tgggtaccgg ggctgaagct ttcgacaaat cttaaattgt gactttgtgc
     2041 atgaaatttt tatatgaacg ttttaaataa gtggacatag tccttcattc ggtatatgca
     2101 gatccggcat caaacacttt ttcaagaaag tttgagtcaa aaggattaaa ggataaagta
     2161 aaacaacaat aacatctccg tgaagcattg agggaagggt ttaactccaa caatgtcact
     2221 gaacaacata acattctgtg tctcacagga tctggatgtt cccctgaaag tgaaaatcaa
     2281 gtcattggaa ggacataagc cactaattgt atccttacag gtatttgata aagagagaaa
     2341 cagaaattta actcttccga tatacacgcc gtatattcct ttcagaaact ctagaacctg
     2401 ggattattgg ttgactttgc ccatacgtat caaacagctt acctttagca gtcatttacg
     2461 cattattttg tgggaataca atggatccaa gcaaattccc tttttcaatc tagaaacgag
     2521 catctttaac ttaaaagact gtactttaaa aagagggttt gaatccttaa aatttcgtta
     2581 tgatgtgatt gaccactgtg aagtggtcac tgataacaaa gatcaagaaa atttgaataa
     2641 atattttcag ggtgaattca cgagactccc ttggcttgac gaaatcacta taagcaaatt
     2701 aaggaaacaa cgggaaaata ggacttggcc tcagggcacc tttgtcttaa acttagaatt
     2761 tccaatgtta gagcttcctg ttgtgttcat cgaaagggaa attatgaata ctcaaatgaa
     2821 tattccaact ttgaagaaca atcctggact aagcacagac ttacgagaac cgaataggaa
     2881 tgatcctcaa atcaaaattt ctttggggga caaatatcac tccacattga agttttacga
     2941 tcctgaccaa ccaaacaatg atccaataga ggaaaagtat aggagattgg aaagagcatc
     3001 taaaaatgca aacttggaca agcaagtgaa gccggacatt aaaaagaggg actacttgaa
     3061 taaaatcatc aactaccccc ctggcactaa attaacagca catgaaaagg gttcaatatg
     3121 gaaatatagg tattatttga tgaacaataa aaaagctctt acaaagttat tacagagtac
     3181 aaatttgaga gaagaatcgg aaagagtaga ggttttagaa ctaatggact catgggcaga
     3241 aattgatata gacgacgcat tagaactatt aggctcaact tttaaaaacc tttcagtgag
     3301 atcgtacgcc gtaaataggc tgaagaaggc atcagacaag gagctggaat tgtacttatt
     3361 acagttggta gaagctgtct gttttgagaa cctgtccact ttttccgaca aatctaacag
     3421 tgaattcact attgtggatg ccgtatcttc gcaaaagctt tcgggtgatt ctatgttact
     3481 atctacatcg catgccaacc aaaaattatt aaagtccatc tcgagtgaat cggaaacttc
     3541 cgggacagaa tcgctaccaa tcgtcatttc tcccttggct gagtttttga ttaggagagc
     3601 actggtcaac ccaaggttag gaagtttttt ctattggtat ttaaagtccg agtctgagga
     3661 caagccatat ttagaccaga ttttaagttc cttttggagt agactggata aaaaatctcg
     3721 gaatatactg aacgatcaag tcaggctaat aaatgtgctc agggaatgtt gtgaaacaat
     3781 taagaggctg aaagatacta cagccaaaaa gatggaatta ctagtgcatt tattggagac
     3841 gaaagtcagg cctcttgtca aagtacgacc aattgcttta ccactagacc ctgatgtgtt
     3901 gatatgcgac gtttgtcctg aaacttcgaa ggtatttaaa agctctttat cgccactaaa
     3961 aataacattc aaaacgacct taaaccaacc atatcactta atgtttaagg ttggcgatga
     4021 tttgagacaa gatcaactgg tagtacagat cataagttta atgaatgagt tattaaaaaa
     4081 tgaaaacgtg gacttgaaat tgacaccata caaaattttg gcaacaggac cacaagaagg
     4141 tgccattgaa tttatcccta atgacacatt agctagcata ctaagcaaat atcacggtat
     4201 tcttggctac cttaaactcc actatcctga tgagaacgcg acattaggcg tacaaggctg
     4261 ggttttagat aattttgtca aatcttgtgc tggctattgt gttattacat acatcttagg
     4321 tgtcggcgat aggcatttag acaacttact agtcacgcca gatgggcact tttttcacgc
     4381 agactttggt tatatcttgg gtcaggaccc caaacctttt ccgccattaa tgaaattgcc
     4441 ccctcaaatt atagaggcgt ttgggggtgc agaatcatca aattatgata aatttcgcag
     4501 ctactgtttt gtggcatatt cgattttaag aaggaatgca ggcttaatct taaacctgtt
     4561 tgaattaatg aagacttcga acatacctga tatcagaata gatcccaatg gtgctatatt
     4621 acgtgtaaga gagagattta atttgaatat gtccgaagaa gatgccacag tgcattttca
     4681 gaatctaatc aatgatagtg taaatgcttt gctgcctatc gtgattgatc atttacataa
     4741 tctggcacaa tactggcgga ccgtcgagga ctacaaagac catgacggtg attataaaga
     4801 tcatgatatc gattacaagg atgacgatga caagtagaat tcgtgagctc gcgaatttct
     4861 tatgatttat gatttttatt attaaataag ttataaaaaa aataagtgta tacaaatttt
     4921 aaagtgactc ttaggtttta aaacgaaaat tcttattctt gagtaactct ttcctgtagg
     4981 tcaggttgct ttctcaggta tagcatgagg tcgctcttat tgaccacacc tctaccggca
     5041 tgccgagcaa atgcctgcaa atcgctcccc gcggccgcca ccgcggtgga gctccagctt
     5101 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     5161 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5221 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5281 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5341 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5401 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5461 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5521 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5581 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5641 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5701 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5761 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5821 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5881 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5941 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     6001 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6061 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6121 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6181 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6241 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6301 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6361 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6421 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6481 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6541 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6601 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6661 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6721 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6781 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6841 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6901 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6961 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     7021 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     7081 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     7141 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     7201 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7261 aggggttccg cgcacatttc cccgaaaagt gccacctggg tccttttcat cacgtgctat
     7321 aaaaataatt ataatttaaa ttttttaata taaatatata aattaaaaat agaaagtaaa
     7381 aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga tgtaaaagac tctaggggga
     7441 tcgccaacaa atactacctt ttatcttgct cttcctgctc tcaggtatta atgccgaatt
     7501 gtttcatctt gtctgtgtag aagaccacac acgaaaatcc tgtgatttta cattttactt
     7561 atcgttaatc gaatgtatat ctatttaatc tgcttttctt gtctaataaa tatatatgta
     7621 aagtacgctt tttgttgaaa ttttttaaac ctttgtttat ttttttttct tcattccgta
     7681 actcttctac cttctttatt tactttctaa aatccaaata caaaacataa aaataaataa
     7741 acacagagta aattcccaaa ttattccatc attaaaagat acgaggcgcg tgtaagttac
     7801 aggcaagcga tccgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     7861 tatcacgagg ccctttcgtc
//
