LOCUS       sequence-164274-        7077 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integrating plasmid, GFP(S65T) under GPD promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7077)
  AUTHORS   Nakanishi K, Weinberg DE, Bartel DP, Patel DJ
  TITLE     Structure of yeast Argonaute with guide RNA.
  JOURNAL   Nature. 2012 Jun 20;486(7403):368-74. doi: 10.1038/nature11211.
  PUBMED    22722195
REFERENCE   2  (bases 1 to 7077)
  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"; journalName: 
            "Nature"; date: "2012-06-20- 20"; volume: "486"; issue: "7403"; 
            pages: "368-74"
COMMENT     Sequence Label: pRS402-GPD-GFP(S65T)
FEATURES             Location/Qualifiers
     source          1..7077
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /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(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /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     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2241..2261
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2243..2261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2286..2929
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2823..2846
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             2947..3663
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2980..3008)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3603..3623
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      3673..3920
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3689..3707)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(3938..3957)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3939..3957)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3967..3984)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3967..3983)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3976..3998)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4128..4583
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4215..4234)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4425..4446
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4788..6503)
                     /codon_start=1
                     /gene="S. cerevisiae ADE2"
                     /product="phosphoribosylaminoimidazole carboxylase, 
                     required for adenine biosynthesis"
                     /label=ADE2
                     /note="yeast auxotrophic marker"
                     /translation="MDSRTVGILGGGQLGRMIVEAANRLNIKTVILDAENSPAKQISNS
                     NDHVNGSFSNPLDIEKLAEKCDVLTIEIEHVDVPTLKNLQVKHPKLKIYPSPETIRLIQ
                     DKYIQKEHLIKNGIAVTQSVPVEQASETSLLNVGRDLGFPFVLKSRTLAYDGRGNFVVK
                     NKEMIPEALEVLKDRPLYAEKWAPFTKELAVMIVRSVNGLVFSYPIVETIHKDNICDLC
                     YAPARVPDSVQLKAKLLAENAIKSFPGCGIFGVEMFYLETGELLINEIAPRPHNSGHYT
                     IDACVTSQFEAHLRSILDLPMPKNFTSFSTITTNAIMLNVLGDKHTKDKELETCERALA
                     TPGSSVYLYGKESRPNRKVGHINIIASSMAECEQRLNYITGRTDIPIKISVAQKLDLEA
                     MVKPLVGIIMGSDSDLPVMSAACAVLKDFGVPFEVTIVSAHRTPHRMSAYAISASKRGI
                     KTIIAGAGGAAHLPGMVAAMTPLPVIGVPVKGSCLDGVDSLHSIVQMPRGVPVATVAIN
                     NSTNAALLAVRLLGAYDSSYTTKMEQFLLKQEEEVLVKAQKLETVGYEAYLENK"
     promoter        complement(6504..6879)
                     /gene="S. cerevisiae ADE2"
                     /label=ADE2 promoter
     primer_bind     complement(6908..6927)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7027..7049
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttac ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcctat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctca
     2281 gtttatcatt atcaatactc gccatttcaa agaatacgta aataattaat agtagtgatt
     2341 ttcctaactt tatttagtca aaaaattagc cttttaattc tgctgtaacc cgtacatgcc
     2401 caaaataggg ggcgggttac acagaatata taacatcgta ggtgtctggg tgaacagttt
     2461 attcctggca tccactaaat ataatggagc ccgcttttta agctggcatc cagaaaaaaa
     2521 aagaatccca gcaccaaaat attgttttct tcaccaacca tcagttcata ggtccattct
     2581 cttagcgcaa ctacagagaa caggggcaca aacaggcaaa aaacgggcac aacctcaatg
     2641 gagtgatgca acctgcctgg agtaaatgat gacacaaggc aattgaccca cgcatgtatc
     2701 tatctcattt tcttacacct tctattacct tctgctctct ctgatttgga aaaagctgaa
     2761 aaaaaaggtt gaaaccagtt ccctgaaatt attcccctac ttgactaata agtatataaa
     2821 gacggtaggt attgattgta attctgtaaa tctatttctt aaacttctta aattctactt
     2881 ttatagttag tctttttttt agttttaaaa caccagaact tagtttcgac ggattctaga
     2941 actagtatgg ttagtaaagg agaagaactt ttcactggag ttgtcccaat tcttgttgaa
     3001 ttagatggtg atgttaatgg gcacaaattt tctgtcagtg gagagggtga aggtgatgca
     3061 acatacggaa aacttaccct taaatttatt tgcactactg gaaaactacc tgttccatgg
     3121 ccaacacttg tcactacttt cacttatggt gttcaatgct tttcaagata cccagatcat
     3181 atgaaacggc atgacttttt caagagtgcc atgcccgaag gttatgtaca ggaaagaact
     3241 atatttttca aagatgacgg gaactacaag acacgtgctg aagtcaagtt tgaaggtgat
     3301 acccttgtta atagaatcga gttaaaaggt attgatttta aagaagatgg aaacattctt
     3361 ggacacaaat tggaatacaa ctataactca cacaatgtat acatcatggc agacaaacaa
     3421 aagaatggaa tcaaagttaa cttcaaaatt agacacaaca ttgaagatgg aagcgttcaa
     3481 ctagcagacc attatcaaca aaatactcca attggcgatg gccctgtcct tttaccagac
     3541 aaccattacc tgtccacaca atctgccctt tcgaaagatc ccaacgaaaa gagagaccac
     3601 atggtccttc ttgagtttgt aacagctgct gggattacac atggcatgga tgaactatac
     3661 aaatagctcg agtcatgtaa ttagttatgt cacgcttaca ttcacgccct ccccccacat
     3721 ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct atttattttt
     3781 ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt ttttttctgt
     3841 acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag gttttgggac
     3901 gctcgaaggc tttaatttgc ggccggtacc caattcgccc tatagtgagt cgtattacgc
     3961 gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     4021 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     4081 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcgacg cgccctgtag
     4141 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     4201 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     4261 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     4321 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     4381 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     4441 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     4501 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     4561 caaaatatta acgtttacaa tttcctgatg cggtattttc tccttacgca tctgtgcggt
     4621 atttcacacc gcatagatct tatgtatgaa attcttaaaa aaggacacct gtaagcgttg
     4681 atttctatgt atgaagtcca catttgatgt aatcataaca aagcctaaaa aataggtata
     4741 tcattttata attatttgct gtacaagtat atcaataaac ttatatatta cttgttttct
     4801 agataagctt cgtaaccgac agtttctaac ttttgtgctt tgacaagaac ttcttcttct
     4861 tgctttaata aaaactgttc cattttcgtt gtataacttg aatcataagc gccaagcagt
     4921 ctgacagcca acagcgcagc gttcgtacta ttattaatag cgacggtagc tactggaaca
     4981 cctctaggca tttgcacaat tgaatgtaaa gaatctactc catctagaca agaacctttt
     5041 acgggcacac cgatgacagg aagtggtgtc attgcagcca ccatacctgg caagtgagca
     5101 gccccaccag ctccagcgat aattgtttta attccacgct tgcttgcgga aatagcatat
     5161 gctgacatcc tatgtggagt tctatgagca gagactattg tcacttcaaa tggaacgcca
     5221 aaatctttta aaaccgcaca tgcggcagac attaccggca agtcagagtc tgatcccatg
     5281 atgattccaa ccaatggttt gaccattgct tccaagtcca acttttgagc gacagagatt
     5341 ttgattggaa tatcagttct acctgtaatg tagttcagcc tttgttcaca ttccgccata
     5401 ctggaggcaa taatatttat gtgacctact tttctgttag gtctagactc ttttccatat
     5461 aagtacactg aggaacctgg agtcgccaat gctctttcgc aagtttctag ctctttatct
     5521 tttgtatgtt tgtctccaag aacatttagc ataatggcgt tcgttgtaat ggtggagaaa
     5581 gatgtgaaat tctttggcat tggcaaatcc aatattgatc tcaaatgagc ttcaaattga
     5641 gaagtgacgc aagcatcaat ggtataatgt ccagagttgt gaggccttgg ggcaatttcg
     5701 ttaataagca attcccctgt ttctaaatag aacatttcca caccaaatat accacaaccg
     5761 ggaaaagatt tgattgcatt ttctgccaac aacttcgcct taagttgaac ggagtccgga
     5821 actctagcag gcgcataaca taagtcacaa atattgtcct tgtggatagt ctctacaatt
     5881 gggtaagaaa acactaaacc gttaacagac ctcacaatca tgactgctaa ttctttagta
     5941 aatggtgccc atttttcggc gtacaaagga cgatccttca gtacttccaa agcttccgga
     6001 atcatttcct tattctttac aacgaagtta cctcttccat cgtatgccaa agtcctcgac
     6061 ttcaagacga atggaaaacc caaatctctt ccaacattca atagggacgt ctcactggct
     6121 tgttccacag gaacactttg ggtaactgct ataccatttt tgattaaatg ctctttttga
     6181 atatatttgt cttgtatcaa tctgattgtt tctggagaag ggtaaatttt taatttggga
     6241 tgttttactt gaagattctt tagtgtagga acatcaacat gctcaatctc aatcgttagc
     6301 acatcacatt tttcagctag tttttcgata tcaagaggat tggaaaagga gccattaacg
     6361 tggtcattgg agttgcttat ttgtttggca ggagaatttt cagcatctag tattaccgtc
     6421 ttaatgttga gcctgtttgc tgcctcaaca atcatacgtc ccaattgtcc ccctcctaat
     6481 ataccaactg ttctagaatc catacttgat tgttttgtcc gattttcttg tttttcttga
     6541 ttgttatagt aggatgtact tagaagagag atccaacgat tttacgcacc aatttataca
     6601 tgaaatgctc cataatattg tccatttagt tcttaataaa aggtcagcaa gagtcaatca
     6661 cttagtatta cccggttcgt agccatgcaa caagagtcat ttgtcagcat agctgtaata
     6721 atcaatcatg acgtaagaaa tgtatcataa ttaaaagttg ttaaagatgt cagtgttatg
     6781 ttggtgttac aaaattctcg gcttctcact aatatttaat atctcttaaa ttttatctgt
     6841 ctttgattct tttaagaaaa gttatgtatt attcaagaat taattcagat ctatggtgca
     6901 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
     6961 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     7021 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcga
//
