LOCUS       sequence-321921-        7940 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gal-inducible yeast expression of cytosolic DHFR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7940)
  AUTHORS   Boos F, Kramer L, Groh C, Jung F, Haberkant P, Stein F, Wollweber F,
            Gackstatter A, Zoller E, van der Laan M, Savitski MM, Benes V, 
            Herrmann JM
  TITLE     Mitochondrial protein-induced stress triggers a global adaptive 
            transcriptional programme.
  JOURNAL   Nat Cell Biol. 2019 Apr;21(4):442-451. doi: 
            10.1038/s41556-019-0294-5. Epub 2019 Mar 18.
  PUBMED    30886345
REFERENCE   2  (bases 1 to 7940)
  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: 
            "10.1038/s41556-019-0294-5"; journalName: "Nat Cell Biol"; date: 
            "2019-04"; volume: "21"; issue: "4"; pages: "442-451"
COMMENT     Sequence Label: pYX233-DHFR
FEATURES             Location/Qualifiers
     source          1..7940
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..665
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    217..334
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     629..652
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             680..1243
                     /codon_start=1
                     /gene="Mus musculus Dhfr"
                     /product="mouse dihydrofolate reductase"
                     /label=DHFR
                     /translation="MVRPLNCIVAVSQNMGIGKNGDLPWPPLRNEFKYFQRMTTTSSVE
                     GKQNLVIMGRKTWFSIPEKNRPLKDRINIVLSRELKEPPRGAHFLAKSLDDALRLIEQP
                     ELASKVDMVWIVGGSSVYQEAMNQPGHLRLFVTRIMQEFESDTFFPEIDLGKYKLLPEY
                     PGVLSEVQEEKGIKYKFEVYEKKD"
     CDS             1265..1291
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1419..1438)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1420..1438)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1445..1462)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1445..1461)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1454..1476)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1602..2057
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1689..1708)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1899..1920
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2160..2834)
                     /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        complement(2835..3116)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3137..3156)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3256..3278
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3316..3334)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     3396..3412
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3396..3412
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     rep_origin      3972..5314
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(4935..4982)
                     /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)."
     promoter        5591..5695
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5696..5764,5765..6556)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFVAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5914..5933)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6727..7315
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7216..7235
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7469..7486
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7603..7624
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7639..7644,7645..7662,7663..7669)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7677..7693
                     /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     7682..7704
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7701..7717
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7701..7717
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        7738..7756
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 tttcaaaaat tcttactttt tttttggatg gacgcaaaga agtttaataa tcatattaca
       61 tggcattacc accatataca tatccatata catatccata tctaatctta cttatatgtt
      121 gtggaaatgt aaagagcccc attatcttag cctaaaaaaa ccttctcttt ggaactttca
      181 gtaatacgct taactgctca ttgctatatt gaagtacgga ttagaagccg ccgagcgggt
      241 gacagccctc cgaaggaaga ctctcctccg tgcgtcctcg tcttcaccgg tcgcgttcct
      301 gaaacgcaga tgtgcctcgc gccgcactgc tccgaacaat aaagattcta caatactagc
      361 ttttatggtt atgaagagga aaaattggca gtaacctggc cccacaaacc ttcaaatgaa
      421 cgaatcaaat taacaaccat aggatgataa tgcgattagt tttttagcct tatttctggg
      481 gtaattaatc agcgaagcga tgatttttga tctattaaca gatatataaa tgcaaaaact
      541 gcataaccac tttaactaat actttcaaca ttttcggttt gtattacttc ttattcaaat
      601 gtaataaaag tatcaacaaa aaattgttaa tatacctcta tactttaacg tcaaggagaa
      661 aaaaccccgg atcgaattca tggttcgacc attgaactgc atcgtcgccg tgtcccaaaa
      721 tatggggatt ggcaagaacg gagacctacc ctggcctccg ctcaggaacg agttcaagta
      781 cttccaaaga atgaccacaa cctcttcagt ggaaggtaaa cagaatctgg tgattatggg
      841 taggaaaacc tggttctcca ttcctgagaa gaatcgacct ttaaaggaca gaattaatat
      901 agttctcagt agagaactca aagaaccacc acgaggagct cattttcttg ccaaaagttt
      961 ggatgatgcc ttaagactta ttgaacaacc ggaattggca agtaaagtag acatggtttg
     1021 gatagtcgga ggcagttctg tttaccagga agccatgaat caaccaggcc acctcagact
     1081 ctttgtgaca aggatcatgc aggaatttga aagtgacacg tttttcccag aaattgattt
     1141 ggggaaatat aaacttctcc cagaataccc aggcgtcctc tctgaggtcc aggaggaaaa
     1201 aggcatcaag tataagtttg aagtctacga gaagaaagac taaaagctta cgcgtcgacc
     1261 cgggtatccg tatgatgtgc ctgactacgc atgatatctc gagctcagct agctaactga
     1321 ataaggaaca atgaacgttt ttcctttctc ttgttcctag tattaatgac tgaccgatac
     1381 atcccttttt ttttttgtct ttgtctagct ccaattcgcc ctatagtgag tcgtattaca
     1441 attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     1501 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     1561 atcgcccttc ccaacagttg cgcagcctga atggcgaatg gacgcgccct gtagcggcgc
     1621 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1681 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1741 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     1801 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1861 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     1921 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     1981 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     2041 attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     2101 caccgcatag atcggcaagt gcacaaacaa tacttaaata aatactactc agtaataacc
     2161 tatttcttag catttttgac gaaatttgct attttgttag agtcttttac accatttgtc
     2221 tccacacctc cgcttacatc aacaccaata acgccattta atctaagcgc atcaccaaca
     2281 ttttctggcg tcagtccacc agctaacata aaatgtaaac tttcggggct ctcttgcctt
     2341 ccaacccagt cagaaatcga gttccaatcc aaaagttcac ctgtcccacc tgcttctgaa
     2401 tcaaacaagg gaataaacga atgaggtttc tgtgaagctg cactgagtag tatgttgcag
     2461 tcttttggaa atacgagtct tttaataact ggcaaaccga ggaactcttg gtattcttgc
     2521 cacgactcat ctccatgcag ttggactata tcaatgccgt aatcattgac cagagccaaa
     2581 acatcctcct taggttgatt acgaaacacg ccaaccaagt atttcggagt gcctgaacta
     2641 tttttatatg cttttacaag acttgaaatt ttccttgcaa taaccgggtc aattgttctc
     2701 tttctattgg gcacacatat aatacccagc aagtcagcat cggaatctag agcacattct
     2761 gcggcctctg tgctctgcaa gccgcaaact ttcaccaatg gaccagaact acctgtgaaa
     2821 ttaataacag acatactcca agctgccttt gtgtgcttaa tcacgtatac tcacgtgctc
     2881 aatagtcacc aatgccctcc ctcttggccc tctccttttc ttttttcgac cgaattaatt
     2941 cttaatcggc aaaaaaagaa aagctccgga tcaagattgt acgtaaggtg acaagctatt
     3001 tttcaataaa gaatatcttc cactactgcc atctggcgtc ataactgcaa agtacacata
     3061 tattacgatg ctgttctatt aaatgcttcc tatattatat atatagtaat gtcgttgatc
     3121 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc
     3181 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     3241 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     3301 gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     3361 tggtttctta gacgtactag tgagagtgcg gccgctctag aactagtgga tcaattccac
     3421 ggactataga ctatactagt atactccgtc tactgtacga tacacttccg ctcaggtcct
     3481 tgtcctttaa cgaggcctta ccactctttt gttactctat tgatccagct cagcaaaggc
     3541 agtgtgatct aagattctat cttcgcgatg tagtaaaact agctagaccg agaaagagac
     3601 tagaaatgca aaaggcactt ctacaatggc tgccatcatt attatccgat gtgacgctgc
     3661 agcttctcaa tgatattcga atacgctttg aggagataca gcctaatatc cgacaaactg
     3721 ttttacagat ttacgatcgt acttgttacc catcattgaa ttttgaacat ccgaacctgg
     3781 gagttttccc tgaaacagat agtatatttg aacctgtata ataatatata gtctagcgct
     3841 ttacggaaga caatgtatgt atttcggttc ctggagaaac tattgcatct attgcatagg
     3901 taatcttgca cgtcgcatcc ccggttcatt ttctgcgttt ccatcttgca cttcaatagc
     3961 atatctttgt taacgaagca tctgtgcttc attttgtaga acaaaaatgc aacgcgagag
     4021 cgctaatttt tcaaacaaag aatctgagct gcatttttac agaacagaaa tgcaacgcga
     4081 aagcgctatt ttaccaacga agaatctgtg cttcattttt gtaaaacaaa aatgcaacgc
     4141 gagagcgcta atttttcaaa caaagaatct gagctgcatt tttacagaac agaaatgcaa
     4201 cgcgagagcg ctattttacc aacaaagaat ctatacttct tttttgttct acaaaaatgc
     4261 atcccgagag cgctattttt ctaacaaagc atcttagatt actttttttc tcctttgtgc
     4321 gctctataat gcagtctctt gataactttt tgcactgtag gtccgttaag gttagaagaa
     4381 ggctactttg gtgtctattt tctcttccat aaaaaaagcc tgactccact tcccgcgttt
     4441 actgattact agcgaagctg cgggtgcatt ttttcaagat aaaggcatcc ccgattatat
     4501 tctataccga tgtggattgc gcatactttg tgaacagaaa gtgatagcgt tgatgattct
     4561 tcattggtca gaaaattatg aacggtttct tctattttgt ctctatatac tacgtatagg
     4621 aaatgtttac attttcgtat tgttttcgat tcactctatg aatagttctt actacaattt
     4681 ttttgtctaa agagtaatac tagagataaa cataaaaaat gtagaggtcg agtttagatg
     4741 caagttcaag gagcgaaagg tggatgggta ggttatatag ggatatagca cagagatata
     4801 tagcaaagag atacttttga gcaatgtttg tggaagcggt attcgcaata ttttagtagc
     4861 tcgttacagt ccggtgcgtt tttggttttt tgaaagtgcg tcttcagagc gcttttggtt
     4921 ttcaaaagcg ctctgaagtt cctatacttt ctagagaata ggaacttcgg aataggaact
     4981 tcaaagcgtt tccgaaaacg agcgcttccg aaaatgcaac gcgagctgcg cacatacagc
     5041 tcactgttca cgtcgcacct atatctgcgt gttgcctgta tatatatata catgagaaga
     5101 acggcatagt gcgtgtttat gcttaaatgc gtacttatat gcgtctattt atgtaggatg
     5161 aaaggtagtc tagtacctcc tgtgatatta tcccattcca tgcggggtat cgtatgcttc
     5221 cttcagcact accctttagc tgttctatat gctgccactc ctcaattgga ttagtctcat
     5281 ccttcaatgc tatcatttcc tttgatattg gatcatatgc atagtaccga gaaactagtg
     5341 cgaagtagtg atcaggtatt gctgttatct gatgagtata cgttgtcctg gccacggcag
     5401 aagcacgctt atcgctccaa tttcccacaa cattagtcaa ctccgttagg cccttcattg
     5461 aaagaaatga ggtcatcaaa tgtcttccaa tgtgagattt tgggccattt tttatagcaa
     5521 agattgaata aggcgcattt ttcttcaaag ctgcggccgc acgtcaggtg gcacttttcg
     5581 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     5641 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     5701 tattcaacat ttccgtgtcg cccttattcc cttttttgtg gcattttgcc ttcctgtttt
     5761 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     5821 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     5881 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     5941 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     6001 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     6061 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     6121 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     6181 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     6241 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     6301 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     6361 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     6421 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     6481 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     6541 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     6601 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     6661 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     6721 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     6781 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     6841 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     6901 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     6961 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     7021 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     7081 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     7141 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     7201 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     7261 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     7321 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     7381 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     7441 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     7501 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     7561 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     7621 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     7681 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agctcgaaat
     7741 taaccctcac taaagggaac aaaagctggt accgggccgg ccgtcggacc aattcgacag
     7801 gttatcagca acaacacagt catatccatt ctcaattagc tctaccacag tgtgtgaacc
     7861 aatgtatcca gcaccacctg taaccaaaac aattttagaa gtactttcac tttgtaactg
     7921 agctgtcatt tatattgaat
//
