LOCUS       sequence-374358-        8219 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression Erk1-GFP-1SIM in yeast cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8219)
  AUTHORS   Sang D, Shu T, Pantoja CF, Ibanez de Opakua A, Zweckstetter M, Holt 
            LJ
  TITLE     Condensed-phase signaling can expand kinase specificity and respond 
            to macromolecular crowding.
  JOURNAL   Mol Cell. 2022 Oct 6;82(19):3693-3711.e10. doi: 
            10.1016/j.molcel.2022.08.016. Epub 2022 Sep 14.
  PUBMED    36108633
REFERENCE   2  (bases 1 to 8219)
  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: 
            "10.1016/j.molcel.2022.08.016"; journalName: "Mol Cell"; date: 
            "2022-10-6- 6"; volume: "82"; issue: "19"; pages: "3693-3711.e10"
COMMENT     Sequence Label: pAV105-PPAB1-GFP-Erk1R84S-1SIM
FEATURES             Location/Qualifiers
     source          1..8219
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     46..65
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(284..388)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     456..474
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     574..593
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(1087..2181)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(2182..2589)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(2890..3345)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3027..3048)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3239..3258
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3471..3493
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3485..3502
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3486..3502
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3512..3531
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3512..3530
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4299..4325
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             4383..5096
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(6442..6462)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(6442..6460)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6481..6497)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6481..6497)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6494..6516)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6505..6521
                     /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)."
     promoter        complement(join(6529..6535,6536..6553,6554..6559))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6574..6595
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6712..6729)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6883..7471)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6963..6982)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7642..8219,1..214,215..283))
                     /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"
ORIGIN
        1 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
       61 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
      121 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
      181 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
      241 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
      301 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
      361 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
      421 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttg
      481 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
      541 gtgttggcgg gtgtcggggc tgccttaact atgcggcatc agagcagatt gtactgagag
      601 tgcaccatat cgactacgtc gttaaggccg tttctgacag agtaaaattc ttgagggaac
      661 tttcaccatt atgggaaatg gttcaagaag gtattgactt aaactccatc aaatggtcag
      721 gtcattgagt gttttttatt tgttgtattt tttttttttt agagaaaatc ctccaatata
      781 taaattagga atcatagttt catgattttc tgttacacct aactttttgt gtggtgccct
      841 cctccttgtc aatattaatg ttaaagtgca attctttttc cttatcacgt tgagccatta
      901 gtatcaattt gcttacctgt attcctttac atcctccttt ttctccttct tgataaatgt
      961 atgtagattg cgtatatagt ttcgtctacc ctatgaacat attccatttt gtaatttcgt
     1021 gtcgtttcta ttatgaattt catttataaa gtttatgtac aaatatcata aaaaaagaga
     1081 atctttttaa gcaaggattt tcttaacttc ttcggcgaca gcatcaccga cttcggtggt
     1141 actgttggaa ccacctaaat caccagttct gatacctgca tccaaaacct ttttaactgc
     1201 atcttcaatg gccttacctt cttcaggcaa gttcaatgac aatttcaaca tcattgcagc
     1261 agacaagata gtggcgatag ggttgacctt attctttggc aaatctggag cagaaccgtg
     1321 gcatggttcg tacaaaccaa atgcggtgtt cttgtctggc aaagaggcca aggacgcaga
     1381 tggcaacaaa cccaaggaac ctgggataac ggaggcttca tcggagatga tatcaccaaa
     1441 catgttgctg gtgattataa taccatttag gtgggttggg ttcttaacta ggatcatggc
     1501 ggcagaatca atcaattgat gttgaacctt caatgtaggg aattcgttct tgatggtttc
     1561 ctccacagtt tttctccata atcttgaaga ggccaaaaca ttagctttat ccaaggacca
     1621 aataggcaat ggtggctcat gttgtagggc catgaaagcg gccattcttg tgattctttg
     1681 cacttctgga acggtgtatt gttcactatc ccaagcgaca ccatcaccat cgtcttcctt
     1741 tctcttacca aagtaaatac ctcccactaa ttctctgaca acaacgaagt cagtaccttt
     1801 agcaaattgt ggcttgattg gagataagtc taaaagagag tcggatgcaa agttacatgg
     1861 tcttaagttg gcgtacaatt gaagttcttt acggattttt agtaaacctt gttcaggtct
     1921 aacactaccg gtaccccatt taggaccacc cacagcacct aacaaaacgg catcagcctt
     1981 cttggaggct tccagcgcct catctggaag tggaacacct gtagcatcga tagcagcacc
     2041 accaattaaa tgattttcga aatcgaactt gacattggaa cgaacatcag aaatagcttt
     2101 aagaacctta atggcttcgg ctgtgatttc ttgaccaacg tggtcacctg gcaaaacgac
     2161 gatcttctta ggggcagaca ttagaatggt atatccttga aatatatata tatatattgc
     2221 tgaaatgtaa aaggtaagaa aagttagaaa gtaagacgat tgctaaccac ctattggaaa
     2281 aaacaatagg tccttaaata atattgtcaa cttcaagtat tgtgatgcaa gcatttagtc
     2341 atgaacgctt ctctattcta tatgaaaagc cggttccggc gctctcacct ttcctttttc
     2401 tcccaatttt tcagttgaaa aaggtatatg cgtcaggcga cctctgaaat taacaaaaaa
     2461 tttccagtca tcgaatttga ttctgtgcga tagcgcccct gtgtgttctc gttatgttga
     2521 ggaaaaaaat aatggttgct aagagattcg aactcttgca tcttacgata cctgagtatt
     2581 cccacagtta actgcggtca agatatttct tgaatcaggc gccttagacc gctcggccaa
     2641 acaaccaatt acttgttgag aaatagagta taattatcct ataaatataa cgtttttgaa
     2701 cacacatgaa caaggaagta caggacaatt gattttgaag agaatgtgga ttttgatgta
     2761 attgttggga ttccattttt aataaggcaa taatattagg tatgtagata tactagaagt
     2821 tctcctcgac cggtcgatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     2881 cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     2941 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     3001 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     3061 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     3121 catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     3181 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     3241 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     3301 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt cgccattcag
     3361 gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc
     3421 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg
     3481 acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac tcactatagg gcgaattggg
     3541 taccgggccc cccctcgaga aggcaagccc agaaaaatat cgcaagcacc tttggtctta
     3601 cagtgccaac ttttggcctg ccgacgttaa gagtacaaag ctgatggcaa tgtacgacaa
     3661 gataacagag tctcaaaaga agtgaaacaa tttttcttca ccacattttc cattgttcct
     3721 tccccccata actataaacg tatttatgta tatatatttg cgtgtaagtg tgtgtactat
     3781 agggcaccgt aaagtaataa tgcttaatta gttactacta tgaccatata agaggtcata
     3841 ctgtatgaag ccacaaagca gatagatcaa tcatgtttaa cgaaaactgt taatcgaaga
     3901 ttatttcttt ttttttttct ctttcctttt tacaaagaaa attttttttg cgctttttgc
     3961 catcaccatc gcaagttctg ggacaattgt tctctttcgc tccagttcca aggaaagagg
     4021 tttctgtttt acttaataga aagtgtcatc ttgtatttta tatctcttct ttcttgtgta
     4081 aaattcttta gttttgattt tgtattttta ggacagtgag ctacgaagta acatttttac
     4141 ttaataaccg tttgaagcat agagcaggcc ctggtatcac cacctaatat ctggcttttt
     4201 attcaataaa aactcaaaaa aaaaaatcca aaaaaaacta aaaaaccaat aaaaataaac
     4261 atatgttgat cacaagtgga gcgtcagggg cacttcagtt gccaccgttg gaaagactaa
     4321 ctctgggctc cgctggatca gctttagcct tgaaattagc aggtcttgat atcctgatca
     4381 ctatgcgtaa aggcgaagag ctgttcactg gtgtcgtccc tattctggtg gaactggatg
     4441 gtgatgtcaa cggtcataag ttttccgtgc gtggcgaggg tgaaggtgac gcaactaatg
     4501 gtaaactgac gctgaagttc atctgtacta ctggtaaact gccggttcct tggccgactc
     4561 tggtaacgac gctgacttat ggtgttcagt gctttgctcg ttatccggac catatgaagc
     4621 agcatgactt cttcaagtcc gccatgccgg aaggctatgt gcaggaacgc acgatttcct
     4681 ttaaggatga cggcacgtac aaaacgcgtg cggaagtgaa atttgaaggc gataccctgg
     4741 taaaccgcat tgagctgaaa ggcattgact ttaaagaaga cggcaatatc ctgggccata
     4801 agctggaata caattttaac agccacaatg tttacatcac cgccgataaa caaaaaaatg
     4861 gcattaaagc gaattttaaa attcgccaca acgtggagga tggcagcgtg cagctggctg
     4921 atcactacca gcaaaacact ccaatcggtg atggtcctgt tctgctgcca gacaatcact
     4981 atctgagcac gcaaagcaaa ctgtctaaag atccgaacga gaaacgcgat catatggttc
     5041 tgctggagtt cgtaaccgca gcgggcatca cgcatggtat ggatgaactg tacaaaggat
     5101 ccggcggaag cggtggctct ggcgggagcg gcggctcgat ggcggcggcg gcggctcagg
     5161 ggggcggggg cggggagccc cgtagaaccg agggggtcgg cccgggggtc ccgggggagg
     5221 tggagatggt gaaggggcag ccgttcgacg tgggcccgcg ctacacgcag ttgcagtaca
     5281 tcggcgaggg cgcgtacggc atggtcagct cggcctatga ccacgtgcgc aagactcgcg
     5341 tggccatcaa gaagatcagc cccttcgaac atcagaccta ctgccagagc acgctccggg
     5401 agatccagat cctgctgcgc ttccgccatg agaatgtcat cggcatccga gacattctgc
     5461 gggcgtccac cctggaagcc atgagagatg tctacattgt gcaggacctg atggagactg
     5521 acctgtacaa gttgctgaaa agccagcagc tgagcaatga ccatatctgc tacttcctct
     5581 accagatcct gcggggcctc aagtacatcc actccgccaa cgtgctccac cgagatctaa
     5641 agccctccaa cctgctcatc aacaccacct gcgaccttaa gatttgtgat ttcggcctgg
     5701 cccggattgc cgatcctgag catgaccaca ccggcttcct gacggagtat gtggctacgc
     5761 gctggtaccg ggccccagag atcatgctga actccaaggg ctataccaag tccatcgaca
     5821 tctggtctgt gggctgcatt ctggctgaga tgctctctaa ccggcccatc ttccctggca
     5881 agcactacct ggatcagctc aaccacattc tgggcatcct gggctcccca tcccaggagg
     5941 acctgaattg tatcatcaac atgaaggccc gaaactacct acagtctctg ccctccaaga
     6001 ccaaggtggc ttgggccaag cttttcccca agtcagactc caaagccctt gacctgctgg
     6061 accggatgtt aacctttaac cccaataaac ggatcacagt ggaggaagcg ctggctcacc
     6121 cctacctgga gcagtactat gacccgacgg atgagccagt ggccgaggag cccttcacct
     6181 tcgccatgga gctggatgac ctacctaagg agcggctgaa ggagctcatc ttccaggaga
     6241 cagcacgctt ccagcccgga gtgctggagg cccccactag aggcggctcg ggaggttcgg
     6301 gcggcagcgg cggaagcaaa gtcgatgtaa ttgacttaac gatcgaatct agcagcgatg
     6361 aagaagaaga tccaccggct aaacgttagt taattaatat ataactgaat aaattttttc
     6421 aaagagctcc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg
     6481 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     6541 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     6601 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     6661 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     6721 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     6781 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     6841 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6901 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6961 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     7021 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     7081 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     7141 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     7201 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     7261 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     7321 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     7381 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     7441 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     7501 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     7561 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     7621 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     7681 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     7741 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagatccac gctcaccggc
     7801 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     7861 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     7921 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     7981 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     8041 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     8101 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     8161 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaat
//
