LOCUS       sequence-138500-        7408 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tryptophan selection; Expression of PP7-2x-yeGFP fusion protein 
            under the MET25 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7408)
  AUTHORS   Saroufim MA, Bensidoun P, Raymond P, Rahman S, Krause MR, Oeffinger 
            M, Zenklusen D
  TITLE     The nuclear basket mediates perinuclear mRNA scanning in budding 
            yeast.
  JOURNAL   J Cell Biol. 2015 Dec 21;211(6):1131-40. doi: 10.1083/jcb.201503070.
  PUBMED    26694838
REFERENCE   2  (bases 1 to 7408)
  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: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2015-12-21- 21"; volume: "211"; 
            issue: "6"; pages: "1131-40"
COMMENT     Sequence Label: pDZ529 pTRP MET25p PP7-PS-2x-yeGFP
FEATURES             Location/Qualifiers
     source          1..7408
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     110..132
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     124..141
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     125..141
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     148..167
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        148..166
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        213..562
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     CDS             968..1678
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             1688..2401
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     terminator      2411..2658
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2427..2445)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(2680..2698)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2719..2735)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2719..2735)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2732..2754)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2743..2759
                     /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(2767..2773,2774..2791,2792..2797))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2812..2833
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    3573..4735
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN4 centromere fused to the 
                     autonomously replicating sequence ARS1/ARS416"
     rep_origin      3898..4735
                     /label=ARS1
                     /note="S. cerevisiae autonomously replicating sequence 
                     ARS1/ARS416"
     CDS             complement(4574..5248)
                     /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(5249..5396)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5416..5434)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5502..5606
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5607..5675,5676..6467)
                     /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(5825..5844)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6638..7226
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7127..7146
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7380..7397
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggcg
       61 aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga
      121 cgttgtaaaa cgacggccag tgaattgtaa tacgactcac tatagggcga attggagctc
      181 cggatgcaag ggttcgaatc ccttagctct cattattttt tgctttttct cttgaggtca
      241 catgatcgca aaatggcaaa tggcacgtga agctgtcgat attggggaac tgtggtggtt
      301 ggcaaatgac taattaagtt agtcaaggcg ccatcctcat gaaaactgtg taacataata
      361 accgaagtgt cgaaaaggtg gcaccttgtc caattgaaca cgctcgatga aaaaaataag
      421 atatatataa ggttaagtaa agcgtctgtt agaaaggaag tttttccttt ttcttgctct
      481 cttgtctttt catctactat ttccttcgtg taatacaggg tcgtcagata catagataca
      541 attctattac ccccatccat actctagaat gtccaaaacc atcgttcttt cggtcggcga
      601 ggctactcgc actctgactg agatccagtc caccgcagac cgtcagatct tcgaagagaa
      661 ggtcgggcct ctggtgggtc ggctgcgcct cacggcttcg ctccgtcaaa acggagccaa
      721 gaccgcgtat cgcgtcaacc taaaactgga tcaggcggac gtcgttgatt ccggacttcc
      781 gaaagtgcgc tacactcagg tatggtcgca cgacgtgaca atcgttgcga atagcaccga
      841 ggcctcgcgc aaatcgttgt acgatttgac caagtccctc gtcgcgacct cgcaggtcga
      901 agatcttgtc gtcaaccttg tgccgctggg ccgtggatcc cccgggctgc aggaattcga
      961 tatcgtgtct aaaggtgaag aattattcac tggtgttgtc ccaattttgg ttgaattaga
     1021 tggtgatgtt aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg atgctactta
     1081 cggtaaattg accttaaaat ttatttgtac tactggtaaa ttgccagttc catggccaac
     1141 cttagtcact actttcggtt atggtgttca atgttttgcg agatacccag atcatatgaa
     1201 acaacatgac tttttcaagt ctgccatgcc agaaggttat gttcaagaaa gaactatttt
     1261 tttcaaagat gacggtaact acaagaccag agctgaagtc aagtttgaag gtgatacctt
     1321 agttaataga atcgaattaa aaggtattga ttttaaagaa gatggtaaca ttttaggtca
     1381 caaattggaa tacaactata actctcacaa tgtttacatc atggctgaca aacaaaagaa
     1441 tggtatcaaa gttaacttca aaattagaca caacattgaa gatggttctg ttcaattagc
     1501 tgaccattat caacaaaata ctccaattgg tgatggtcca gtcttgttac cagacaacca
     1561 ttacttatcc actcaatctg ccttatccaa agatccaaac gaaaagagag accacatggt
     1621 cttgttagaa tttgttactg ctgctggtat tacccatggt atggatgaat tgtacaaaga
     1681 tatcgtgtct aaaggtgaag aattattcac tggtgttgtc ccaattttgg ttgaattaga
     1741 tggtgatgtt aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg atgctactta
     1801 cggtaaattg accttaaaat ttatttgtac tactggtaaa ttgccagttc catggccaac
     1861 cttagtcact actttcggtt atggtgttca atgttttgcg agatacccag atcatatgaa
     1921 acaacatgac tttttcaagt ctgccatgcc agaaggttat gttcaagaaa gaactatttt
     1981 tttcaaagat gacggtaact acaagaccag agctgaagtc aagtttgaag gtgatacctt
     2041 agttaataga atcgaattaa aaggtattga ttttaaagaa gatggtaaca ttttaggtca
     2101 caaattggaa tacaactata actctcacaa tgtttacatc atggctgaca aacaaaagaa
     2161 tggtatcaaa gttaacttca aaattagaca caacattgaa gatggttctg ttcaattagc
     2221 tgaccattat caacaaaata ctccaattgg tgatggtcca gtcttgttac cagacaacca
     2281 ttacttatcc actcaatctg ccttatccaa agatccaaac gaaaagagag accacatggt
     2341 cttgttagaa tttgttactg ctgctggtat tacccatggt atggatgaat tgtacaaata
     2401 actggtcgag tcatgtaatt agttatgtca cgcttacatt cacgccctcc ccccacatcc
     2461 gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat ttattttttt
     2521 atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac
     2581 agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt tttgggacgc
     2641 tcgaaggctt taatttgcgg ccggtaccag cttttgttcc ctttagtgag ggttaatttc
     2701 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     2761 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     2821 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     2881 ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
     2941 tgaatggcga atggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3001 accgcatata tcgctgggcc attctcatga agaatatctt gaatttattg tcatattact
     3061 agttggtgtg gaagtccata tatcggtgat caatatagtg gttgacatgc tggctagtca
     3121 acattgagcc ttttgatcat gcaaatatat tacggtattt tacaatcaaa tatcaaactt
     3181 aactattgac tttataactt atttaggtgg taacattctt ataaaaaaga aaaaaattac
     3241 tgcaaaacag tactagcttt taacttgtat cctaggttat ctatgctgtc tcaccataga
     3301 gaatattacc tatttcagaa tgtatgtcca tgattcgccg ggtaaataca tataatacac
     3361 aaatctggct taataaagtc tataatatat ctcataaaga agtgctaaat tggctagtgc
     3421 tatatatttt taagaaaatt tcttttgact aagtccatat cgactttgta aaagttcact
     3481 ttagcataca tatattacac gagccagaaa ttgtaacttt tgcctaaaat cacaaattgc
     3541 aaaatttaat tgcttgcaaa aggtcacatg cttataatca acttttttaa aaatttaaaa
     3601 tactttttta ttttttattt ttaaacataa atgaaataat ttatttattg tttatgatta
     3661 ccgaaacata aaacctgctc aagaaaaaga aactgttttg tccttggaaa aaaagcacta
     3721 cctaggagcg gccaaaatgc cgaggctttc atagcttaaa ctctttacag aaaataggca
     3781 ttatagatca gttcgagttt tcttattctt ccttccggtt ttatcgtcac agttttacag
     3841 taaataagta tcacctctta gagttcgatg ataagctgtc aaacatgaga attaattcca
     3901 catgttaaaa tagtgaagga gcatgttcgg cacacagtgg accgaacgtg gggtaagtgc
     3961 actagggtcc ggttaaacgg atctcgcatt gatgaggcaa cgctaattat caacatatag
     4021 attgttatct atctgcatga acacgaaatc tttacttgac gacttgaggc tgatggtgtt
     4081 tatgcaaaga aaccactgtg tttaatatgt gtcactgttt gatattactg tcagcgtaga
     4141 agataatagt aaaagcggtt aataagtgta tttgagataa gtgtgataaa gtttttacag
     4201 cgaaaagacg ataaatacaa gaaaatgatt acgaggatac ggagagaggt atgtacatgt
     4261 gtatttatat actaagctgc cggcggttgt ttgcaagacc gagaaaaggc tagcaagaat
     4321 cgggtcattg tagcgtatgc gcctgtgaac attctcttca acaagtttga ttccattgcg
     4381 gtgaaatggt aaaagtcaac cccctgcgat gtatattttc ctgtacaatc aatcaaaaag
     4441 ccaaatgatt tagcattatc tttacatctt gttattttac agattttatg tttagatctt
     4501 ttatgcttgc ttttcaaaag gcttgcaggc aagtgcacaa acaatactta aataaatact
     4561 actcagtaat aacctatttc ttagcatttt tgacgaaatt tgctattttg ttagagtctt
     4621 ttacaccatt tgtctccaca cctccgctta catcaacacc aataacgcca tttaatctaa
     4681 gcgcatcacc aacattttct ggcgtcagtc caccagctaa cataaaatgt aagctttcgg
     4741 ggctctcttg ccttccaacc cagtcagaaa tcgagttcca atccaaaagt tcacctgtcc
     4801 cacctgcttc tgaatcaaac aagggaataa acgaatgagg tttctgtgaa gctgcactga
     4861 gtagtatgtt gcagtctttt ggaaatacga gtcttttaat aactggcaaa ccgaggaact
     4921 cttggtattc ttgccacgac tcatctccat gcagttggac gatatcaatg ccgtaatcat
     4981 tgaccagagc caaaacatcc tccttaggtt gattacgaaa cacgccaacc aagtatttcg
     5041 gagtgcctga actattttta tatgctttta caagacttga aattttcctt gcaataaccg
     5101 ggtcaattgt tctctttcta ttgggcacac atataatacc cagcaagtca gcatcggaat
     5161 ctagagcaca ttctgcggcc tctgtgctct gcaagccgca aactttcacc aatggaccag
     5221 aactacctgt gaaattaata acagacatac tccaagctgc ctttgtgtgc ttaatcacgt
     5281 atactcacgt gctcaatagt caccaatgcc ctccctcttg gccctctcct tttctttttt
     5341 cgaccgaatt aattcttaat cggcaaaaaa agaaaagctc cggatcaaga ttgtacaatt
     5401 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     5461 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     5521 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     5581 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     5641 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     5701 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     5761 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     5821 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     5881 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     5941 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     6001 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     6061 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     6121 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     6181 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     6241 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     6301 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     6361 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     6421 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     6481 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     6541 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     6601 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     6661 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     6721 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     6781 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6841 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6901 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6961 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     7021 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     7081 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     7141 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     7201 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     7261 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     7321 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     7381 gcgagtcagt gagcgaggaa gcggaaga
//
