LOCUS       sequence-167611-        8428 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  yeast expression of mutant EEF2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8428)
  AUTHORS   Davydova E, Ho AY, Malecki J, Moen A, Enserink JM, Jakobsson ME, 
            Loenarz C, Falnes PO
  TITLE     Identification and characterization of a novel evolutionarily 
            conserved lysine-specific methyltransferase targeting eukaryotic 
            translation elongation factor 2 (eEF2).
  JOURNAL   J Biol Chem. 2014 Oct 31;289(44):30499-510. doi: 
            10.1074/jbc.M114.601658. Epub 2014 Sep 17.
  PUBMED    25231979
REFERENCE   2  (bases 1 to 8428)
  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"; doi: 
            "10.1074/jbc.M114.601658"; journalName: "J Biol Chem"; date: 
            "2014-10-31- 31"; volume: "289"; issue: "44"; pages: "30499-510"
COMMENT     Sequence Label: pYES260-sc.eEF2 K509A
FEATURES             Location/Qualifiers
     source          1..8428
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(31..486)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     380..399
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        505..947
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    505..622
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     918..941
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     979..998
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        979..997
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1014..1031
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1038..1058
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      3684..3931
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3700..3718)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4001..4018)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4172..4760)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4252..4271)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4931..5722,5723..5791))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLID*MEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(5887..6690)
                     /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"
     promoter        complement(6691..6911)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      7510..8390
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 aaaacaggaa gattgtataa gcaaatattt aaattgtaaa cgttaatatt ttgttaaaat
       61 tcgcgttaaa tttttgttaa atcagctcat tttttaacga atagcccgaa atcggcaaaa
      121 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttccaaca
      181 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaagg gtctatcagg
      241 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
      301 aagcagtaaa tcggaagggt aaacggatgc ccccatttag agcttgacgg ggaaagccgg
      361 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggggctagg gcggtgggaa
      421 gtgtaggggt cacgctgggc gtaaccacca cacccgccgc gcttaatggg gcgctacagg
      481 gcgcgtgggg atgatccact agtacggatt agaagccgcc gagcgggtga cagccctccg
      541 aaggaagact ctcctccgtg cgtcctcgtc ctcaccggtc gcgttcctga aacgcagatg
      601 tgcctcgcgc cgcactgctc cgaacaataa agattctaca atactagctt ttatggttat
      661 gaagaggaaa aattggcagt aacctggccc cacaaacctt caaatgaacg aatcaaatta
      721 acaaccatag gatgataatg cgattagttt tttagcctta tttctggggt aattaatcag
      781 cgaagcgatg atttttgatc tattaacaga tatataaatg caaaaactgc attaaccact
      841 ttaactaata ctttcaacat tttcggtttg tattacttct tattcaaatg taataaaagt
      901 atcaacaaaa aattgttaat atacctctat actttaacgt caaggagaaa aaaccccgga
      961 tcggactact agcagctgta atacgactca ctatagggaa tattaagctt atgcaccatc
     1021 accatcacca tgctagcgag aatctttatt ttcagggcgc catggaggtt gctttcactg
     1081 ttgaccaaat gcgttcttta atggacaaag ttaccaatgt gcgtaacatg tccgttattg
     1141 ctcacgtcga tcatggtaag tccactttga ccgattcctt ggtccaaaga gccggtatta
     1201 tttccgctgc taaggctggt gaagctcgtt tcaccgatac cagaaaggat gaacaagaaa
     1261 gaggtatcac tatcaagtct accgctattt ctctatactc tgaaatgtct gacgaagatg
     1321 tcaaggaaat caagcaaaag accgacggta actccttctt gatcaacttg atcgactctc
     1381 caggtcacgt tgacttctcc tctgaagtta ctgccgcttt acgtgtcact gacggtgctt
     1441 tggttgtcgt cgacaccatt gaaggtgtct gtgtccaaac cgaaactgtt ttgagacaag
     1501 ctttgggtga gagaatcaag cctgttgttg ttatcaacaa ggtcgacaga gctttgttgg
     1561 aattgcaagt ttctaaggaa gatttatacc aaacctttgc cagaactgtt gaatccgtta
     1621 acgtcatcgt ttccacctac gccgatgaag ttttgggtga tgtccaagtt tacccagcca
     1681 gaggtaccgt tgccttcggt tccggtttgc acggttgggc tttcactatc cgtcaattcg
     1741 ccaccagata tgctaagaaa ttcggtgtcg acaaggccaa gatgatggac agattatggg
     1801 gtgactcttt cttcaaccca aagaccaaga agtggaccaa caaggacact gatgctgaag
     1861 gtaagccatt ggaaagagct ttcaacatgt tcatcttgga cccaatcttc agattattca
     1921 ctgctatcat gaacttcaag aaagatgaaa ttccagtttt gctagaaaag ttggaaattg
     1981 tcttgaaggg tgacgaaaag gacttggaag gtaaggcctt gttgaaggtt gttatgagaa
     2041 agttcttgcc agctgccgat gccttattgg aaatgattgt cttgcacttg ccatctccag
     2101 tcactgctca agcctacaga gctgaacaat tatacgaagg tccagctgac gatgccaact
     2161 gtattgctat caagaactgt gatccaaagg ctgatttgat gttgtacgtc tccaagatgg
     2221 tgccaacctc tgataagggt agattctacg ccttcggtag agtttttgcc ggtactgtta
     2281 agtccggtca aaaggtcaga atccaaggtc caaactacgt tccaggtaag aaggacgatt
     2341 tgttcatcaa ggccattcaa agagttgttt tgatgatggg tagatttgtc gaaccaatcg
     2401 atgactgtcc agccggtaac attatcggtt tagtcggtat cgatcaattc ttgttgaaga
     2461 ctggtacttt gaccaccagt gaaactgctc acaacatgaa ggtcatgaaa ttctctgtct
     2521 ctccagttgt gcaagtcgct gtcgaagtca agaacgctaa cgacttacca aaattggtcg
     2581 aaggtttggc gagattgtcc aagtctgatc catgtgtctt gacctatatg tctgaatccg
     2641 gtgaacatat cgttgctggt accggtgaat tgcatttgga aatttgtttg caagatttgg
     2701 aacacgacca cgctggtgtt ccattgaaga tctccccacc agttgtcgct tacagagaaa
     2761 ctgttgaaag tgaatcttct caaactgctt tgtccaagtc tccaaacaag cataacagaa
     2821 tctacttgaa ggctgaacca attgacgaag aagtctcttt ggctattgaa aacggtatca
     2881 tcaacccaag agatgatttc aaggccagag ctagaatcat ggctgacgac tacggttggg
     2941 atgtcaccga tgccagaaag atctggtgtt tcggtccaga cggtaacggt ccaaacttgg
     3001 ttattgacca aactaaggct gtccaatact tgcacgaaat caaggattcc gttgttgctg
     3061 ctttccaatg ggctaccaag gaaggtccaa ttttcggtga agaaatgaga tctgtcagag
     3121 ttaacatttt ggatgttact ttacatgccg atgctatcca cagaggtggt ggtcaaatca
     3181 tcccaaccat gagaagagct acttacgccg gtttcttgtt ggctgatcca aagatccaag
     3241 aaccagtttt cttggtcgaa attcaatgtc cagaacaagc cgtcggtggt atctactccg
     3301 tcttaaacaa gaagagaggt caagtcgttt ctgaagaaca aagaccaggt actccattgt
     3361 ttaccgtcaa ggcctacttg ccagttaacg aatctttcgg tttcactggt gaattgagac
     3421 aagctaccgg tggtcaagct ttcccacaaa tggttttcga ccattggtcc actttaggtt
     3481 ctgacccatt ggacccaacc tctaaggctg gtgaaattgt tcttgctgct cgtaagagac
     3541 acggtatgaa ggaagaagtt ccaggctggc aagaatatta cgacaaattg taaggatcca
     3601 ctagtaacgg ccgccagtgt gctggaattc tgcagatatc catcacactg gcggccgctc
     3661 gagcatgcat ctagagggcc gcatcatgta attagttatg tcacgcttac attcacgccc
     3721 tccccccaca tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc
     3781 tatttatttt tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct
     3841 tttttttctg tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa
     3901 ggttttggga cgctcgaagg ctttaatttg cggccctgca ttaatgaatc ggccaacgcg
     3961 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4021 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4081 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaagccca
     4141 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4201 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4261 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4321 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4381 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4441 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4501 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4561 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4621 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4681 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4741 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4801 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     4861 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4921 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4981 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagcgcttac
     5041 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5101 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5161 cctccattca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5221 gtttgcgcaa cgttgttggc attgctacag gcatcgtggt gtcactctcg tcgtttggta
     5281 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5341 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5401 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5461 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5521 gaccgagttg ctcttgcccg gcgtcaatac gggataatag tgtatcacat agcagaactt
     5581 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5641 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5701 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5761 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatgg gtaataactg
     5821 atataattaa attgaagctc taatttgtga gtttagtata catgcattta cttataatac
     5881 agttttttag ttttgctggc cgcatcttct caaatatgct tcccagcctg cttttctgta
     5941 acgttcaccc tctaccttag catcccttcc ctttgcaaat agtcctcttc caacaataat
     6001 aatgtcagat cctgtagaga ccacatcatc cacggttcta tactgttgac ccaatgcgtc
     6061 tcccttgtca tctaaaccca caccgggtgt cataatcaac caatcgtaac cttcatctct
     6121 tccacccatg tctctttgag caataaagcc gataacaaaa tctttgtcgc tcttcgcaat
     6181 gtcaacagta cccttagtat attctccagt agatagggag cccttgcatg acaattctgc
     6241 taacatcaaa aggcctctag gttcctttgt tacttcttct gccgcctgct tcaaaccgct
     6301 aacaatacct gggcccacca caccgtgtgc attcgtaatg tctgcccatt ctgctattct
     6361 gtatacaccc gcagagtact gcaatttgac tgtattacca atgtcagcaa attttctgtc
     6421 ttcgaagagt aaaaaattgt acttggcgga taatgccttt agcggcttaa ctgtgccctc
     6481 cattgaaaaa tcagtcaaga tatccacatg tgtttttagt aaacaaattt tgggacctaa
     6541 tgcttcaact aactccagta attccttggt ggtacgaaca tccaatgaag cacacaagtt
     6601 tgtttgcttt tcgtgcatga tattaaatag cttggcagca acaggactag gatgagtagc
     6661 agcacgttcc ttatatgtag ctttcgacat gatttatctt cgtttcctgc aggtttttgt
     6721 tctgtgcagt tgggttaaga atactgggca atttcatgtt tcttcaacac tacatatgcg
     6781 tatatatacc aatctaagtc tgtgctcctt ccttcgttct tccttctgtt cggagattac
     6841 cgaatcaaaa aaatttcaaa gaaaccgaaa tcaaaaaaaa gaataaaaaa aaaatgatga
     6901 attgaattga aaagctagct tatcgatgat aagctgtcaa agatgagaat taattccacg
     6961 gactatagac tatactagat actccgtcta ctgtacgata cacttccgct caggtccttg
     7021 tcctttaacg aggccttacc actcttttgt tactctattg atccagctca gcaaaggcag
     7081 tgtgatctaa gattctatct tcgcgatgta gtaaaactag ctagaccgag aaagagacta
     7141 gaaatgcaaa aggcacttct acaatggctg ccatcattat tatccgatgt gacgctgcag
     7201 cttctcaatg atattcgaat acgctttgag gagatacagc ctaatatccg acaaactgtt
     7261 ttacagattt acgatcgtac ttgttaccca tcattgaatt ttgaacatcc gaacctggga
     7321 gttttccctg aaacagatag tatatttgaa cctgtataat aatatatagt ctagcgcttt
     7381 acggaagaca atgtatgtat ttcggttcct ggagaaacta ttgcatctat tgcataggta
     7441 atcttgcacg tcgcatcccc ggttcatttt ctgcgtttcc atcttgcact tcaatagcat
     7501 atctttgtta acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa cgcgagagcg
     7561 ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaaa
     7621 gcgctatttt accaacgaag aatctgtgct tcatttttgt aaaacaaaaa tgcaacgcga
     7681 cgagagcgct aatttttcaa acaaagaatc tgagctgcat ttttacagaa cagaaatgca
     7741 acgcgagagc gctattttac caacaaagaa tctatacttc ttttttgttc tacaaaaatg
     7801 catcccgaga gcgctatttt tctaacaaag catcttagat tacttttttt ctcctttgtg
     7861 cgctctataa tgcagtctct tgataacttt ttgcactgta ggtccgttaa ggttagaaga
     7921 aggctacttt ggtgtctatt ttctcttcca taaaaaaagc ctgactccac ttcccgcgtt
     7981 tactgattac tagcgaagct gcgggtgcat tttttcaaga taaaggcatc cccgattata
     8041 ttctataccg atgtggattg cgcatacttt gtgaacagaa agtgatagcg ttgatgattc
     8101 ttcattggtc agaaaattat gaacggtttc ttctattttg tctctatata ctacgtatag
     8161 gaaatgttta cattttcgta ttgttttcga ttcactctat gaatagttct tactacaatt
     8221 tttttgtcta aagagtaata ctagagataa acataaaaaa tgtagaggtc gagtttagat
     8281 gcaagttcaa ggagcgaaag gtggatgggt aggttatata gggatatagc acagagatat
     8341 atagcaaaga gatacttttg agcaatgttt gtggaagcgg tattcgcaat gggaagctcc
     8401 accccggttg ataatcagaa aagcccca
//
