LOCUS       sequence-169277-        7235 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 7235)
  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 7235)
  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..7235
                     /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"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     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      4124..4579
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4211..4230)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4421..4442
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4784..6499)
                     /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(6500..6875)
                     /gene="S. cerevisiae ADE2"
                     /label=ADE2 promoter
     primer_bind     complement(7066..7085)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7185..7207
                     /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 aatactgttc
     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 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcgtat 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 tggacgcgcc ctgtagcggc
     4141 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     4201 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     4261 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4321 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4381 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4441 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     4501 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4561 atattaacgc ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4621 cacaccgcat agatcttatg tatgaaattc ttaaaaaagg acacctgtaa gcgttgattt
     4681 ctatgtatga agtccacatt tgatgtaatc ataacaaagc ctaaaaaata ggtatatcat
     4741 tttataatta tttgctgtac aagtatatca ataaacttat atattacttg ttttctagat
     4801 aagcttcgta accgacagtt tctaactttt gtgctttgac aagaacttct tcttcttgct
     4861 ttaataaaaa ctgttccatt ttcgttgtat aacttgaatc ataagcgcca agcagtctga
     4921 cagccaacag cgcagcgttc gtactattat taatagcgac ggtagctact ggaacacctc
     4981 taggcatttg cacaattgaa tgtaaagaat ctactccatc tagacaagaa ccttttacgg
     5041 gcacaccgat gacaggaagt ggtgtcattg cagccaccat acctggcaag tgagcagccc
     5101 caccagctcc agcgataatt gttttaattc cacgcttgct tgcggaaata gcatatgctg
     5161 acatcctatg tggagttcta tgagcagaga ctattgtcac ttcaaatgga acgccaaaat
     5221 cttttaaaac cgcacatgcg gcagacatta ccggcaagtc agagtctgat cccatgatga
     5281 ttccaaccaa tggtttgacc attgcttcca agtccaactt ttgagcgaca gagattttga
     5341 ttggaatatc agttctacct gtaatgtagt tcagcctttg ttcacattcc gccatactgg
     5401 aggcaataat atttatgtga cctacttttc tgttaggtct agactctttt ccatataagt
     5461 acactgagga acctggagtc gccaatgctc tttcgcaagt ttctagctct ttatcttttg
     5521 tatgtttgtc tccaagaaca tttagcataa tggcgttcgt tgtaatggtg gagaaagatg
     5581 tgaaattctt tggcattggc aaatccaata ttgatctcaa atgagcttca aattgagaag
     5641 tgacgcaagc atcaatggta taatgtccag agttgtgagg ccttggggca atttcgttaa
     5701 taagcaattc ccctgtttct aaatagaaca tttccacacc aaatatacca caaccgggaa
     5761 aagatttgat tgcattttct gccaacaact tcgccttaag ttgaacggag tccggaactc
     5821 tagcaggcgc ataacataag tcacaaatat tgtccttgtg gatagtctct acaattgggt
     5881 aagaaaacac taaaccgtta acagacctca caatcatgac tgctaattct ttagtaaatg
     5941 gtgcccattt ttcggcgtac aaaggacgat ccttcagtac ttccaaagct tccggaatca
     6001 tttccttatt ctttacaacg aagttacctc ttccatcgta tgccaaagtc ctcgacttca
     6061 agacgaatgg aaaacccaaa tctcttccaa cattcaatag ggacgtctca ctggcttgtt
     6121 ccacaggaac actttgggta actgctatac catttttgat taaatgctct ttttgaatat
     6181 atttgtcttg tatcaatctg attgtttctg gagaagggta aatttttaat ttgggatgtt
     6241 ttacttgaag attctttagt gtaggaacat caacatgctc aatctcaatc gttagcacat
     6301 cacatttttc agctagtttt tcgatatcaa gaggattgga aaaggagcca ttaacgtggt
     6361 cattggagtt gcttatttgt ttggcaggag aattttcagc atctagtatt accgtcttaa
     6421 tgttgagcct gtttgctgcc tcaacaatca tacgtcccaa ttgtccccct cctaatatac
     6481 caactgttct agaatccata cttgattgtt ttgtccgatt ttcttgtttt tcttgattgt
     6541 tatagtagga tgtacttaga agagagatcc aacgatttta cgcaccaatt tatacatgaa
     6601 atgctccata atattgtcca tttagttctt aataaaaggt cagcaagagt caatcactta
     6661 gtattacccg gttcgtagcc atgcaacaag agtcatttgt cagcatagct gtaataatca
     6721 atcatgacgt aagaaatgta tcataattaa aagttgttaa agatgtcagt gttatgttgg
     6781 tgttacaaaa ttctcggctt ctcactaata tttaatatct cttaaatttt atctgtcttt
     6841 gattctttta agaaaagtta tgtattattc aagaattaat tcagatctat tcttttgatt
     6901 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     6961 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc ctgatgcggt attttctcct
     7021 tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga
     7081 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     7141 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     7201 tcagaggttt tcaccgtcat caccgaaacg cgcga
//
