LOCUS       sequence-322692-        7202 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EGFP expression under the control of the Galactose-inducible 
            promoter in yeast. Contains 2 um element for high copy within 
            yeast..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7202)
  AUTHORS   Harel I, Chen YR, Ziv I, Singh PP, Heinzer D, Navarro Negredo P, 
            Goshtchevsky U, Wang W, Astre G, Moses E, McKay A, Machado BE, 
            Hebestreit K, Yin S, Sanchez Alvarado A, Jarosz DF, Brunet A
  TITLE     Identification of protein aggregates in the aging vertebrate brain 
            with prion-like and phase-separation properties.
  JOURNAL   Cell Rep. 2024 Jun 25;43(6):112787. doi: 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28.
  PUBMED    38810650
REFERENCE   2  (bases 1 to 7202)
  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: "Cell Rep.";
            date: "2024-06-25"; volume: "43(6):112787. doi"; pages: " 
            10.1016/j.celrep.2023.112787. Epub 2024 May 28"
COMMENT     Sequence Label: pAG426GAL-EGFP
FEATURES             Location/Qualifiers
     source          1..7202
                     /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"
     CDS             join(4149..4151,4152..4154,4155..4868)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4149..4151,4152..4154,4155..4865)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4194..4215)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4455..4474)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4802..4823
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4873..4897)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(4930..4946)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4931..4947)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      4949..5196
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4965..4983)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(5214..5233)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5215..5233)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5243..5260)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5243..5259)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5252..5274)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5404..5859
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5491..5510)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5701..5722
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5990..6793)
                     /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(6794..7009)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(7033..7052)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7152..7174
                     /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 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     4201 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     4261 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     4321 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     4381 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     4441 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     4501 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     4561 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     4621 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     4681 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     4741 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     4801 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     4861 acaagtaata agacccagct ttcttgtaca aagtggtgat gggctgcagg aattcgatat
     4921 caagcttatc gataccgtcg acctcgagtc atgtaattag ttatgtcacg cttacattca
     4981 cgccctcccc ccacatccgc tctaaccgaa aaggaaggag ttagacaacc tgaagtctag
     5041 gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat atttcaaatt
     5101 tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa aaccttgctt
     5161 gagaaggttt tgggacgctc gaaggcttta atttgcggcc ggtacccaat tcgccctata
     5221 gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     5281 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     5341 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc
     5401 gcgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     5461 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5521 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5581 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     5641 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5701 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     5761 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     5821 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttc ctgatgcggt attttctcct
     5881 tacgcatctg tgcggtattt cacaccgcat agggtaataa ctgatataat taaattgaag
     5941 ctctaatttg tgagtttagt atacatgcat ttacttataa tacagttttt tagttttgct
     6001 ggccgcatct tctcaaatat gcttcccagc ctgcttttct gtaacgttca ccctctacct
     6061 tagcatccct tccctttgca aatagtcctc ttccaacaat aataatgtca gatcctgtag
     6121 agaccacatc atccacggtt ctatactgtt gacccaatgc gtctcccttg tcatctaaac
     6181 ccacaccggg tgtcataatc aaccaatcgt aaccttcatc tcttccaccc atgtctcttt
     6241 gagcaataaa gccgataaca aaatctttgt cgctcttcgc aatgtcaaca gtacccttag
     6301 tatattctcc agtagatagg gagcccttgc atgacaattc tgctaacatc aaaaggcctc
     6361 taggttcctt tgttacttct tctgccgcct gcttcaaacc gctaacaata cctgggccca
     6421 ccacaccgtg tgcattcgta atgtctgccc attctgctat tctgtataca cccgcagagt
     6481 actgcaattt gactgtatta ccaatgtcag caaattttct gtcttcgaag agtaaaaaat
     6541 tgtacttggc ggataatgcc tttagcggct taactgtgcc ctccatggaa aaatcagtca
     6601 agatatccac atgtgttttt agtaaacaaa ttttgggacc taatgcttca actaactcca
     6661 gtaattcctt ggtggtacga acatccaatg aagcacacaa gtttgtttgc ttttcgtgca
     6721 tgatattaaa tagcttggca gcaacaggac taggatgagt agcagcacgt tccttatatg
     6781 tagctttcga catgatttat cttcgtttcc tgcaggtttt tgttctgtgc agttgggtta
     6841 agaatactgg gcaatttcat gtttcttcaa cactacatat gcgtatatat accaatctaa
     6901 gtctgtgctc cttccttcgt tcttccttct gttcggagat taccgaatca aaaaaatttc
     6961 aaggaaaccg aaatcaaaaa aaagaataaa aaaaaaatga tgaattgaaa aggtggtatg
     7021 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
     7081 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     7141 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     7201 ga
//
