LOCUS       sequence-466838-        8446 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Galactose inducible expression of EWSR1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8446)
  AUTHORS   Chavez Lab
  TITLE     Multiplex screening identifies broad modifiers of ALS-associated 
            proteotoxicity
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 8446)
  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: 
            "unpublished"
COMMENT     Sequence Label: pAG426-GAL-EWSR1
FEATURES             Location/Qualifiers
     source          1..8446
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      69..1411
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    401..448
                     /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        1438..1542
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1543..1611,1612..2403)
                     /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(1761..1780)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2574..3162
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3063..3082
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3316..3333
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3450..3471
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3486..3491,3492..3509,3510..3516)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3524..3540
                     /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     3529..3551
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3548..3564
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3548..3564
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3583..3603
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3585..3603
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3627..4068
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3627..3744
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     4039..4062
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     4083..4099
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4083..4099
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    4107..4131
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(6121..6145)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(6178..6194)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6179..6195)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      6197..6444
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(6213..6231)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6462..6481)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6463..6481)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6491..6508)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6491..6507)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6500..6522)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6648..7103
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6735..6754)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6945..6966
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(7234..8037)
                     /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(8038..8253)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(8277..8296)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8396..8418
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagtatga tccaatatca aaggaaatga tagcattgaa ggatgagact aatccaattg
      121 aggagtggca gcatatagaa cagctaaagg gtagtgctga aggaagcata cgataccccg
      181 catggaatgg gataatatca caggaggtac tagactacct ttcatcctac ataaatagac
      241 gcatataagt acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata
      301 tatacaggca acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc
      361 gcgttgcatt ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt
      421 cctattctct agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
      481 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
      541 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
      601 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
      661 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
      721 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
      781 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
      841 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
      901 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
      961 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     1021 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     1081 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     1141 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     1201 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     1261 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     1321 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     1381 gttctacaaa atgaagcaca gatgcttcgt tcaggtggca cttttcgggg aaatgtgcgc
     1441 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1501 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     1561 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     1621 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     1681 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     1741 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     1801 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     1861 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     1921 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     1981 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     2041 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     2101 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     2161 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     2221 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     2281 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     2341 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     2401 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     2461 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     2521 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     2581 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     2641 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     2701 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     2761 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     2821 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     2881 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     2941 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     3001 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     3061 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     3121 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     3181 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     3241 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     3301 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     3361 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     3421 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     3481 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     3541 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
     3601 agggaacaaa agctggagct ctagtacgga ttagaagccg ccgagcgggt gacagccctc
     3661 cgaaggaaga ctctcctccg tgcgtcctcg tcttcaccgg tcgcgttcct gaaacgcaga
     3721 tgtgcctcgc gccgcactgc tccgaacaat aaagattcta caatactagc ttttatggtt
     3781 atgaagagga aaaattggca gtaacctggc cccacaaacc ttcaaatgaa cgaatcaaat
     3841 taacaaccat aggatgataa tgcgattagt tttttagcct tatttctggg gtaattaatc
     3901 agcgaagcga tgatttttga tctattaaca gatatataaa tgcaaaaact gcataaccac
     3961 tttaactaat actttcaaca ttttcggttt gtattacttc ttattcaaat gtaataaaag
     4021 tatcaacaaa aaattgttaa tatacctcta tactttaacg tcaaggagaa aaaaccccgg
     4081 attctagaac tagtggatcc cccatcacaa gtttgtacaa aaaagcaggc ttcgaaggag
     4141 ataacaaaat ggcgtccacg gattacagta cctatagcca agctgcagcg cagcagggct
     4201 acagtgctta caccgcccag cccactcaag gatatgcaca gaccacccag gcatatgggc
     4261 aacaaagcta tggaacctat ggacagccca ctgatgtcag ctatacccag gctcagacca
     4321 ctgcaaccta tgggcagacc gcctatgcaa cttcttatgg acagcctccc actggttata
     4381 ctactccaac tgccccccag gcatacagcc agcctgtcca ggggtatggc actggtgctt
     4441 atgataccac cactgctaca gtcaccacca cccaggcctc ctatgcagct cagtctgcat
     4501 atggcactca gcctgcttat ccagcctatg ggcagcagcc agcagccact gcacctacaa
     4561 gaccgcagga tggaaacaag cccactgaga ctagtcaacc tcaatctagc acagggggtt
     4621 acaaccagcc cagcctagga tatggacaga gtaactacag ttatccccag gtacctggga
     4681 gctaccccat gcagccagtc actgcacctc catcctaccc tcctaccagc tattcctcta
     4741 cacagccgac tagttatgat cagagcagtt actctcagca gaacacctat gggcaaccga
     4801 gcagctatgg acagcagagt agctatggtc aacaaagcag ctatgggcag cagcctccca
     4861 ctagttaccc accccaaact ggatcctaca gccaagctcc aagtcaatat agccaacaga
     4921 gcagcagcta cgggcagcag agttcattcc gacaggacca ccccagtagc atgggtgttt
     4981 atgggcagga gtctggagga ttttccggac caggagagaa ccggagcatg agtggccctg
     5041 ataaccgggg caggggaaga gggggatttg atcgtggagg catgagcaga ggtgggcggg
     5101 gaggaggacg cggtggaatg ggcagcgctg gagagcgagg tggcttcaat aagcctggtg
     5161 gacccatgga tgaaggacca gatcttgatc taggcccacc tgtagatcca gatgaagact
     5221 ctgacaacag tgcaatttat gtacaaggat taaatgacag tgtgactcta gatgatctgg
     5281 cagacttctt taagcagtgt ggggttgtta agatgaacaa gagaactggg caacccatga
     5341 tccacatcta cctggacaag gaaacaggaa agcccaaagg cgatgccaca gtgtcctatg
     5401 aagacccacc cactgccaag gctgccgtgg aatggtttga tgggaaagat tttcaaggga
     5461 gcaaacttaa agtctccctt gctcggaaga agcctccaat gaacagtatg cggggtggtc
     5521 tgccaccccg tgagggcaga ggcatgccac caccactccg tggaggtcca ggaggcccag
     5581 gaggtcctgg gggacccatg ggtcgcatgg gaggccgtgg aggagataga ggaggcttcc
     5641 ctccaagagg accccggggt tcccgaggga acccctctgg aggaggaaac gtccagcacc
     5701 gagctggaga ctggcagtgt cccaatccgg gttgtggaaa ccagaacttc gcctggagaa
     5761 cagagtgcaa ccagtgtaag gccccaaagc ctgaaggctt cctcccgcca ccctttccgc
     5821 ccccgggtgg tgatcgtggc agaggtggcc ctggtggcat gcggggagga agaggtggcc
     5881 tcatggatcg tggtggtccc ggtggaatgt tcagaggtgg ccgtggtgga gacagaggtg
     5941 gcttccgtgg tggccggggc atggaccgag gtggctttgg tggaggaaga cgaggtggcc
     6001 ctggggggcc ccctggacct ttgatggaac agatgggagg aagaagagga ggacgtggag
     6061 gacctggaaa aatggataaa ggcgagcacc gtcaggagcg cagagatcgg ccctactagg
     6121 acccagcttt cttgtacaaa gtggtgatgg gctgcaggaa ttcgatatca agcttatcga
     6181 taccgtcgac ctcgagtcat gtaattagtt atgtcacgct tacattcacg ccctcccccc
     6241 acatccgctc taaccgaaaa ggaaggagtt agacaacctg aagtctaggt ccctatttat
     6301 ttttttatag ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt
     6361 ctgtacagac gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga gaaggttttg
     6421 ggacgctcga aggctttaat ttgcggccgg tacccaattc gccctatagt gagtcgtatt
     6481 acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     6541 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     6601 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggacg cgccctgtag
     6661 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     6721 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     6781 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     6841 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     6901 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     6961 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     7021 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     7081 caaaatatta acgcttacaa tttcctgatg cggtattttc tccttacgca tctgtgcggt
     7141 atttcacacc gcatagggta ataactgata taattaaatt gaagctctaa tttgtgagtt
     7201 tagtatacat gcatttactt ataatacagt tttttagttt tgctggccgc atcttctcaa
     7261 atatgcttcc cagcctgctt ttctgtaacg ttcaccctct accttagcat cccttccctt
     7321 tgcaaatagt cctcttccaa caataataat gtcagatcct gtagagacca catcatccac
     7381 ggttctatac tgttgaccca atgcgtctcc cttgtcatct aaacccacac cgggtgtcat
     7441 aatcaaccaa tcgtaacctt catctcttcc acccatgtct ctttgagcaa taaagccgat
     7501 aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga
     7561 tagggagccc ttgcatgaca attctgctaa catcaaaagg cctctaggtt cctttgttac
     7621 ttcttctgcc gcctgcttca aaccgctaac aatacctggg cccaccacac cgtgtgcatt
     7681 cgtaatgtct gcccattctg ctattctgta tacacccgca gagtactgca atttgactgt
     7741 attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa aaattgtact tggcggataa
     7801 tgcctttagc ggcttaactg tgccctccat ggaaaaatca gtcaagatat ccacatgtgt
     7861 ttttagtaaa caaattttgg gacctaatgc ttcaactaac tccagtaatt ccttggtggt
     7921 acgaacatcc aatgaagcac acaagtttgt ttgcttttcg tgcatgatat taaatagctt
     7981 ggcagcaaca ggactaggat gagtagcagc acgttcctta tatgtagctt tcgacatgat
     8041 ttatcttcgt ttcctgcagg tttttgttct gtgcagttgg gttaagaata ctgggcaatt
     8101 tcatgtttct tcaacactac atatgcgtat atataccaat ctaagtctgt gctccttcct
     8161 tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa tttcaaggaa accgaaatca
     8221 aaaaaaagaa taaaaaaaaa atgatgaatt gaaaaggtgg tatggtgcac tctcagtaca
     8281 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     8341 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     8401 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcga
//
