LOCUS       sequence-430950-        6718 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Galactose inducible expression of Kar2-Beta-Amyloid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6718)
  AUTHORS   Chavez Lab
  TITLE     Multiplex screening identifies broad modifiers of ALS-associated 
            proteotoxicity
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 6718)
  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            "unpublished"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-Kar2-Beta-Amyloid
FEATURES             Location/Qualifiers
     source          1..6718
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    26..47
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(62..67,68..85,86..92)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    100..116
                     /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     105..127
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     124..140
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     124..140
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     159..179
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        161..179
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        203..644
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    203..320
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     615..638
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     659..675
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     659..675
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    683..707
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(969..993)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(1026..1042)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1027..1043)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      1045..1292
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1061..1079)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(1310..1329)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1311..1329)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1339..1356)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1339..1355)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1348..1370)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1496..1951
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1583..1602)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1793..1814
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2082..2885)
                     /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(2886..3101)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3125..3144)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3244..3266
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3304..3322)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      3363..4705
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    3695..3742
                     /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        4732..4836
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4837..4905,4906..5697)
                     /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(5055..5074)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5868..6456
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6357..6376
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6610..6627
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg
       61 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
      121 acacaggaaa cagctatgac catgattacg ccaagcgcgc aattaaccct cactaaaggg
      181 aacaaaagct ggagctctag tacggattag aagccgccga gcgggtgaca gccctccgaa
      241 ggaagactct cctccgtgcg tcctcgtctt caccggtcgc gttcctgaaa cgcagatgtg
      301 cctcgcgccg cactgctccg aacaataaag attctacaat actagctttt atggttatga
      361 agaggaaaaa ttggcagtaa cctggcccca caaaccttca aatgaacgaa tcaaattaac
      421 aaccatagga tgataatgcg attagttttt tagccttatt tctggggtaa ttaatcagcg
      481 aagcgatgat ttttgatcta ttaacagata tataaatgca aaaactgcat aaccacttta
      541 actaatactt tcaacatttt cggtttgtat tacttcttat tcaaatgtaa taaaagtatc
      601 aacaaaaaat tgttaatata cctctatact ttaacgtcaa ggagaaaaaa ccccggattc
      661 tagaactagt ggatccccca tcacaagttt gtacaaaaaa gcaggctaca aaatgttttt
      721 caacagacta agcgctggca agctgctggt accactctcc gtggtcctgt acgccctttt
      781 cgtggtaata ttacctttac agaattcttt ccactcctcc aatgttttag ttagaggtga
      841 tgctgaattt agacatgatt ctggttatga agttcatcat caaaaattgg ttttttttgc
      901 tgaagatgtt ggttctaata aaggtgctat tattggtttg atggttggtg gtgttgtcat
      961 tgcttaaaac ccagctttct tgtacaaagt ggtgatgggc tgcaggaatt cgatatcaag
     1021 cttatcgata ccgtcgacct cgagtcatgt aattagttat gtcacgctta cattcacgcc
     1081 ctccccccac atccgctcta accgaaaagg aaggagttag acaacctgaa gtctaggtcc
     1141 ctatttattt ttttatagtt atgttagtat taagaacgtt atttatattt caaatttttc
     1201 ttttttttct gtacagacgc gtgtacgcat gtaacattat actgaaaacc ttgcttgaga
     1261 aggttttggg acgctcgaag gctttaattt gcggccggta cccaattcgc cctatagtga
     1321 gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     1381 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     1441 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg
     1501 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     1561 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     1621 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     1681 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     1741 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     1801 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     1861 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     1921 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     1981 tgtgcggtat ttcacaccgc atagggtaat aactgatata attaaattga agctctaatt
     2041 tgtgagttta gtatacatgc atttacttat aatacagttt tttagttttg ctggccgcat
     2101 cttctcaaat atgcttccca gcctgctttt ctgtaacgtt caccctctac cttagcatcc
     2161 cttccctttg caaatagtcc tcttccaaca ataataatgt cagatcctgt agagaccaca
     2221 tcatccacgg ttctatactg ttgacccaat gcgtctccct tgtcatctaa acccacaccg
     2281 ggtgtcataa tcaaccaatc gtaaccttca tctcttccac ccatgtctct ttgagcaata
     2341 aagccgataa caaaatcttt gtcgctcttc gcaatgtcaa cagtaccctt agtatattct
     2401 ccagtagata gggagccctt gcatgacaat tctgctaaca tcaaaaggcc tctaggttcc
     2461 tttgttactt cttctgccgc ctgcttcaaa ccgctaacaa tacctgggcc caccacaccg
     2521 tgtgcattcg taatgtctgc ccattctgct attctgtata cacccgcaga gtactgcaat
     2581 ttgactgtat taccaatgtc agcaaatttt ctgtcttcga agagtaaaaa attgtacttg
     2641 gcggataatg cctttagcgg cttaactgtg ccctccatgg aaaaatcagt caagatatcc
     2701 acatgtgttt ttagtaaaca aattttggga cctaatgctt caactaactc cagtaattcc
     2761 ttggtggtac gaacatccaa tgaagcacac aagtttgttt gcttttcgtg catgatatta
     2821 aatagcttgg cagcaacagg actaggatga gtagcagcac gttccttata tgtagctttc
     2881 gacatgattt atcttcgttt cctgcaggtt tttgttctgt gcagttgggt taagaatact
     2941 gggcaatttc atgtttcttc aacactacat atgcgtatat ataccaatct aagtctgtgc
     3001 tccttccttc gttcttcctt ctgttcggag attaccgaat caaaaaaatt tcaaggaaac
     3061 cgaaatcaaa aaaaagaata aaaaaaaaat gatgaattga aaaggtggta tggtgcactc
     3121 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     3181 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     3241 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     3301 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttagt
     3361 atgatccaat atcaaaggaa atgatagcat tgaaggatga gactaatcca attgaggagt
     3421 ggcagcatat agaacagcta aagggtagtg ctgaaggaag catacgatac cccgcatgga
     3481 atgggataat atcacaggag gtactagact acctttcatc ctacataaat agacgcatat
     3541 aagtacgcat ttaagcataa acacgcacta tgccgttctt ctcatgtata tatatataca
     3601 ggcaacacgc agatataggt gcgacgtgaa cagtgagctg tatgtgcgca gctcgcgttg
     3661 cattttcgga agcgctcgtt ttcggaaacg ctttgaagtt cctattccga agttcctatt
     3721 ctctagaaag tataggaact tcagagcgct tttgaaaacc aaaagcgctc tgaagacgca
     3781 ctttcaaaaa accaaaaacg caccggactg taacgagcta ctaaaatatt gcgaataccg
     3841 cttccacaaa cattgctcaa aagtatctct ttgctatata tctctgtgct atatccctat
     3901 ataacctacc catccacctt tcgctccttg aacttgcatc taaactcgac ctctacattt
     3961 tttatgttta tctctagtat tactctttag acaaaaaaat tgtagtaaga actattcata
     4021 gagtgaatcg aaaacaatac gaaaatgtaa acatttccta tacgtagtat atagagacaa
     4081 aatagaagaa accgttcata attttctgac caatgaagaa tcatcaacgc tatcactttc
     4141 tgttcacaaa gtatgcgcaa tccacatcgg tatagaatat aatcggggat gcctttatct
     4201 tgaaaaaatg cacccgcagc ttcgctagta atcagtaaac gcgggaagtg gagtcaggct
     4261 ttttttatgg aagagaaaat agacaccaaa gtagccttct tctaacctta acggacctac
     4321 agtgcaaaaa gttatcaaga gactgcatta tagagcgcac aaaggagaaa aaaagtaatc
     4381 taagatgctt tgttagaaaa atagcgctct cgggatgcat ttttgtagaa caaaaaagaa
     4441 gtatagattc tttgttggta aaatagcgct ctcgcgttgc atttctgttc tgtaaaaatg
     4501 cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg ttttacaaaa
     4561 atgaagcaca gattcttcgt tggtaaaata gcgctttcgc gttgcatttc tgttctgtaa
     4621 aaatgcagct cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat ttttgttcta
     4681 caaaatgaag cacagatgct tcgttcaggt ggcacttttc ggggaaatgt gcgcggaacc
     4741 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     4801 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     4861 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     4921 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     4981 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     5041 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     5101 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     5161 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     5221 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     5281 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     5341 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     5401 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     5461 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     5521 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     5581 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     5641 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     5701 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     5761 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     5821 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     5881 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     5941 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     6001 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     6061 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     6121 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     6181 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     6241 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     6301 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     6361 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     6421 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     6481 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     6541 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     6601 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct
     6661 ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc cgactgga
//
