LOCUS       sequence-472852-        8012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression of ZZ-FL yeast Atg14 in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8012)
  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 8012)
  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: pYO411
FEATURES             Location/Qualifiers
     source          1..8012
                     /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        191..472
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             473..1147
                     /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"
     rep_origin      complement(1250..1705)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1387..1408)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1599..1618
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1831..1853
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1845..1862
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1846..1862
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1872..1891
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1872..1890
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1939..1963
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    2021..2138
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2319..2342
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             2475..2495
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2511..2684
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             2685..2858
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             2868..2888
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2919..2939
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2961..2981
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(4410..4430)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4410..4428)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4449..4465)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4449..4465)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4462..4484)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4473..4489
                     /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(4497..4503,4504..4521,4522..4527))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4542..4563
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4680..4697)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4851..5439)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4931..4950)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5610..6401,6402..6470))
                     /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     6233..6252
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6471..6575)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6602..7944
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(7565..7612)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     7985..8003
                     /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 accatagatc aacgacatta ctatatatat aatataggaa gcatttaata gaacagcatc
      241 gtaatatatg tgtactttgc agttatgacg ccagatggca gtagtggaag atattcttta
      301 ttgaaaaata gcttgtcacc ttacgtacaa tcttgatccg gagcttttct ttttttgccg
      361 attaagaatt aattcggtcg aaaaaagaaa aggagagggc caagagggag ggcattggtg
      421 actattgagc acgtgagtat acgtgattaa gcacacaaag gcagcttgga gtatgtctgt
      481 tattaatttc acaggtagtt ctggtccatt ggtgaaagtt tgcggcttgc agagcacaga
      541 ggccgcagaa tgtgctctag attccgatgc tgacttgctg ggtattatat gtgtgcccaa
      601 tagaaagaga acaattgacc cggttattgc aaggaaaatt tcaagtcttg taaaagcata
      661 taaaaatagt tcaggcactc cgaaatactt ggttggcgtg tttcgtaatc aacctaagga
      721 ggatgttttg gctctggtca atgattacgg cattgatatc gtccaactgc atggagatga
      781 gtcgtggcaa gaataccaag agttcctcgg tttgccagtt attaaaagac tcgtatttcc
      841 aaaagactgc aacatactac tcagtgcagc ttcacagaaa cctcattcgt ttattccctt
      901 gtttgattca gaagcaggtg ggacaggtga acttttggat tggaactcga tttctgactg
      961 ggttggaagg caagagagcc ccgaaagctt acattttatg ttagctggtg gactgacgcc
     1021 agaaaatgtt ggtgatgcgc ttagattaaa tggcgttatt ggtgttgatg taagcggagg
     1081 tgtggagaca aatggtgtaa aagactctaa caaaatagca aatttcgtca aaaatgctaa
     1141 gaaataggtt attactgagt agtatttatt taagtattgt ttgtgcactt gccgatctat
     1201 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcagga aattgtaagc
     1261 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     1321 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     1381 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     1441 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     1501 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     1561 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     1621 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     1681 cttaatgcgc cgctacaggg cgcgtccatt cgccattcag gctgcgcaac tgttgggaag
     1741 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     1801 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     1861 gtgagcgcgc gtaatacgac tcactatagg gcgaattggg taccgggccc cccctcgagt
     1921 acttctcgac aagctatcac cactttgtac aagaaagctg ggtctagata tctcgagtgc
     1981 ggccgcggat ctttgcaaag atccaaaaaa agcatgcgta cggattagaa gccgccgagc
     2041 gggtgacagc cctccgaagg aagactctcc tccgtgcgtc ctcgtcttca ccggtcgcgt
     2101 tcctgaaacg cagatgtgcc tcgcgccgca ctgctccgaa caataaagat tctacaatac
     2161 tagcttttat ggttatgaag aggaaaattg gcagtaacct ggccccacaa acctcaaatg
     2221 aacgaatcaa attaacaacc agatatctcg actgaaaaaa aaggtttaaa ccagttccct
     2281 gaaattattc ccctacttga ctaataagta tataaagacg gtaggtattg attgtaattc
     2341 tgtaaatcta tttcttaaac ttcttgaatt ctacttttat agttagtctt ttttttagtt
     2401 ttaaaacacc aagaacttag tttcgaataa acacacataa ataaaccatg atgggagcag
     2461 gggcgggtgc tagcgagaac ctgtacttcc agggcgccca cgatgaagcc gtagacaaca
     2521 aattcaacaa agaacaacaa aacgcgttct atgagatctt acatttacct aacttaaacg
     2581 aagaacaacg aaacgccttc atccaaagtt taaaagatga cccaagccaa agcgctaacc
     2641 ttttagcaga agctaaaaag ctaaatgatg ctcaggcgcc gaaagtagac aacaaattca
     2701 acaaagaaca acaaaacgcg ttctatgaga tcttacattt acctaactta aacgaagaac
     2761 aacgaaacgc cttcatccaa agtttaaaag atgacccaag ccaaagcgct aaccttttag
     2821 cagaagctaa aaagctaaat gatgctcagg cgccgaaagt agacgcggaa aacctgtatt
     2881 tccagggagg cggatcttct agaccaacta ctgcctctga aaatttgtac tttcaaggcg
     2941 agttgaagac cactgcttct gaaaacttgt acttccaagg tgagttactc gagggtgaat
     3001 tgaagactgc tgctttggct caacacgacg aagccatggg catgcattgc ccaatttgcc
     3061 accatagagc gcatgtagtg tactgtgcac attgtatcaa tacgagccca agtctgctac
     3121 tgaagctaaa actagattta attttattaa aggatgagaa taaagaactt aacgggaaag
     3181 tcgaacaaat attaaacgag gccatgaact atgaccagtt agatatcaag aggatggaga
     3241 aaaagaaaga tcccctgatg aatagtctca tgaagttaga tgttttacga atgaaaaaga
     3301 ataataacct gattaggcac aggatagaac agctaaatga gcggatttat agtaaaagga
     3361 atcacatcag cgaattgaaa gtagaaattg acaattataa atgttataag gtgggtactg
     3421 gtacggacaa attaagagag caagtggaaa ttagtgatgc aaaaaataag ctggcacaag
     3481 tatcgaagat ttgtgaatcg gctagagatt acaagctcaa tctactaaat aattggtttg
     3541 tgatccagaa gctgcaagat aatttccaaa tacctttcgc tatagctttt cagccactaa
     3601 tatctttgaa aaacttccgt attttacctc ttgctataac aaacgattcc atcaacatca
     3661 tgtggaagta tattagcttc ttctcagaca ttcttatgat taaacttccc tacacaaata
     3721 aaatctgtga acaacccatg tttgagtttt ccgatagcat acagacagta gtacaaaggt
     3781 tgatcaagct tatcataaat attttacaga tatgtagaca tttgaaactc gtaccttcaa
     3841 cacccatgga tatcccatgg ctactggacc agtacgatgt ggatgggctg ttctataata
     3901 tggtgaaacg gaacaagatg aagtgtaggt ccgtctcgct atattggact tttgggatgt
     3961 tgtactcgat ggttttggat aacatgaata atccacaaag aggacatcct gcaaggcgga
     4021 ccgcaccacc tccaacagtc acaggacctc atgaccgatg gtacgtggta ggcggatcct
     4081 aacccaagct taagtcgact ttgttcccac tgtactttta gctcgtacaa aatacaatat
     4141 acttttcatt tctccgtaaa caacatgttt tcccatgtaa tatccttttc tatttttcgt
     4201 tccgttacca actttacaca tactttatat agctattcac ttctatacac taaaaaacta
     4261 agacaatttt aattttgctg cctgccatat ttcaatttgt tataaattcc tataatttat
     4321 cctattagta gctaaaaaaa gatgaatgtg aatcgaatcc taagagaatt cccctcgaag
     4381 atcgatctgg cgagctccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     4441 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4501 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4561 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4621 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4681 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4741 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4801 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4861 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4921 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4981 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5041 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5101 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5161 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5221 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5281 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5341 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5401 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5461 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5521 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5581 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5641 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5701 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5761 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5821 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5881 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5941 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     6001 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6061 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6121 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6181 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6241 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6301 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6361 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6421 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6481 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6541 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6601 gaacgaagca tctgtgcttc attttgtaga acaaaaatgc aacgcgagag cgctaatttt
     6661 tcaaacaaag aatctgagct gcatttttac agaacagaaa tgcaacgcga aagcgctatt
     6721 ttaccaacga agaatctgtg cttcattttt gtaaaacaaa aatgcaacgc gagagcgcta
     6781 atttttcaaa caaagaatct gagctgcatt tttacagaac agaaatgcaa cgcgagagcg
     6841 ctattttacc aacaaagaat ctatacttct tttttgttct acaaaaatgc atcccgagag
     6901 cgctattttt ctaacaaagc atcttagatt actttttttc tcctttgtgc gctctataat
     6961 gcagtctctt gataactttt tgcactgtag gtccgttaag gttagaagaa ggctactttg
     7021 gtgtctattt tctcttccat aaaaaaagcc tgactccact tcccgcgttt actgattact
     7081 agcgaagctg cgggtgcatt ttttcaagat aaaggcatcc ccgattatat tctataccga
     7141 tgtggattgc gcatactttg tgaacagaaa gtgatagcgt tgatgattct tcattggtca
     7201 gaaaattatg aacggtttct tctattttgt ctctatatac tacgtatagg aaatgtttac
     7261 attttcgtat tgttttcgat tcactctatg aatagttctt actacaattt ttttgtctaa
     7321 agagtaatac tagagataaa cataaaaaat gtagaggtcg agtttagatg caagttcaag
     7381 gagcgaaagg tggatgggta ggttatatag ggatatagca cagagatata tagcaaagag
     7441 atacttttga gcaatgtttg tggaagcggt attcgcaata ttttagtagc tcgttacagt
     7501 ccggtgcgtt tttggttttt tgaaagtgcg tcttcagagc gcttttggtt ttcaaaagcg
     7561 ctctgaagtt cctatacttt ctagagaata ggaacttcgg aataggaact tcaaagcgtt
     7621 tccgaaaacg agcgcttccg aaaatgcaac gcgagctgcg cacatacagc tcactgttca
     7681 cgtcgcacct atatctgcgt gttgcctgta tatatatata catgagaaga acggcatagt
     7741 gcgtgtttat gcttaaatgc gtacttatat gcgtctattt atgtaggatg aaaggtagtc
     7801 tagtacctcc tgtgatatta tcccattcca tgcggggtat cgtatgcttc cttcagcact
     7861 accctttagc tgttctatat gctgccactc ctcaattgga ttagtctcat ccttcaatgc
     7921 tatcatttcc tttgatattg gatcatacta agaaaccatt attatcatga cattaaccta
     7981 taaaaatagg cgtatcacga ggccctttcg tc
//
