LOCUS       sequence-226341-        7322 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP-X2 where the second consensus motif's lysine is mutated to 
            arginine.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7322)
  AUTHORS   Rossl A, Denoncourt A, Lin MS, Downey M
  TITLE     A synthetic non-histone substrate to study substrate targeting by 
            the Gcn5 HAT and sirtuin HDACs.
  JOURNAL   J Biol Chem. 2019 Apr 19;294(16):6227-6239. doi: 
            10.1074/jbc.RA118.006051. Epub 2019 Feb 25.
  PUBMED    30804216
REFERENCE   2  (bases 1 to 7322)
  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: 
            "10.1074/jbc.RA118.006051"; journalName: "J Biol Chem"; date: 
            "2019-04-19- 19"; volume: "294"; issue: "16"; pages: "6227-6239"
COMMENT     Sequence Label: GFP-X2R2
FEATURES             Location/Qualifiers
     source          1..7322
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     14..30
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     14..30
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             50..763
                     /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="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(80..108)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     703..723
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(811..827)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(812..828)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      830..1077
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(846..864)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(1099..1117)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1138..1154)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1138..1154)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1151..1173)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1162..1178
                     /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(1186..1192,1193..1210,1211..1216))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1231..1252
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1369..1386)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1540..2128)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1620..1639)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2299..3090,3091..3159))
                     /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     2922..2941
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3160..3264)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    3301..3804
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     3845..3863
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3901..3923)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4023..4042
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4066..4281
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             4282..5085
                     /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"
     rep_origin      complement(5216..5671)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5353..5374)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5565..5584
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5797..5819
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5811..5828
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5812..5828
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5835..5854
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5835..5853
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        6618..7322
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
ORIGIN
        1 ccaagctggc cgctctagaa ctagtggatc ccccgggctg caggaattca tgagtaaagg
       61 agaagaactt ttcactggag ttgtcccaat tcttgttgaa ttagatggtg atgttaatgg
      121 gcacaaattt tctgtcagtg gagagggtga aggtgatgca acatacggaa aacttaccct
      181 taaatttatt tgcactactg gaaaactacc tgttccatgg ccaacacttg tcactacttt
      241 cacttatggt gttcaatgct tttcaagata cccagatcat atgaaacggc atgacttttt
      301 caagagtgcc atgcccgaag gttatgtaca ggaaagaact atatttttca aagatgacgg
      361 gaactacaag acacgtgctg aagtcaagtt tgaaggtgat acccttgtta atagaatcga
      421 gttaaaaggt attgatttta aagaagatgg aaacattctt ggacacaaat tggaatacaa
      481 ctataactca cacaatgtat acatcatggc agacaaacaa aagaatggaa tcaaagttaa
      541 cttcaaaatt agacacaaca ttgaagatgg aagcgttcaa ctagcagacc attatcaaca
      601 aaatactcca attggcgatg gccctgtcct tttaccagac aaccattacc tgtccacaca
      661 atctgccctt tcgaaagatc ccaacgaaaa gagagaccac atggtccttc ttgagtttgt
      721 aacagctgct gggattacac atggcatgga tgaactatac aaaggttctt caaaaagacc
      781 tggatcatct aggaggcctt aaaagcttat cgataccgtc gacctcgagt catgtaatta
      841 gttatgtcac gcttacattc acgccctccc cccacatccg ctctaaccga aaaggaagga
      901 gttagacaac ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga
      961 acgttattta tatttcaaat ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac
     1021 attatactga aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcggc
     1081 cggtaccagc ttttgttccc tttagtgagg gttaatttcg agcttggcgt aatcatggtc
     1141 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     1201 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     1261 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     1321 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     1381 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     1441 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     1501 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1561 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1621 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     1681 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1741 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     1801 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     1861 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     1921 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     1981 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2041 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2101 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2161 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2221 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2281 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2341 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     2401 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     2461 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     2521 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     2581 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     2641 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     2701 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     2761 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     2821 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     2881 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     2941 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     3001 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     3061 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     3121 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     3181 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     3241 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ggtccttttc
     3301 atcacgtgct ataaaaataa ttataattta aattttttaa tataaatata taaattaaaa
     3361 atagaaagta aaaaaagaaa ttaaagaaaa aatagttttt gttttccgaa gatgtaaaag
     3421 actctagggg gatcgccaac aaatactacc ttttatcttg ctcttcctgc tctcaggtat
     3481 taatgccgaa ttgtttcatc ttgtctgtgt agaagaccac acacgaaaat cctgtgattt
     3541 tacattttac ttatcgttaa tcgaatgtat atctatttaa tctgcttttc ttgtctaata
     3601 aatatatatg taaagtacgc tttttgttga aattttttaa acctttgttt attttttttt
     3661 cttcattccg taactcttct accttcttta tttactttct aaaatccaaa tacaaaacat
     3721 aaaaataaat aaacacagag taaattccca aattattcca tcattaaaag atacgaggcg
     3781 cgtgtaagtt acaggcaagc gatccgtcta agaaaccatt attatcatga cattaaccta
     3841 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     3901 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     3961 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     4021 tgcggcatca gagcagattg tactgagagt gcaccatacc accttttcaa ttcatcattt
     4081 tttttttatt cttttttttg atttcggttt ccttgaaatt tttttgattc ggtaatctcc
     4141 gaacagaagg aagaacgaag gaaggagcac agacttagat tggtatatat acgcatatgt
     4201 agtgttgaag aaacatgaaa ttgcccagta ttcttaaccc aactgcacag aacaaaaacc
     4261 tgcaggaaac gaagataaat catgtcgaaa gctacatata aggaacgtgc tgctactcat
     4321 cctagtcctg ttgctgccaa gctatttaat atcatgcacg aaaagcaaac aaacttgtgt
     4381 gcttcattgg atgttcgtac caccaaggaa ttactggagt tagttgaagc attaggtccc
     4441 aaaatttgtt tactaaaaac acatgtggat atcttgactg atttttccat ggagggcaca
     4501 gttaagccgc taaaggcatt atccgccaag tacaattttt tactcttcga agacagaaaa
     4561 tttgctgaca ttggtaatac agtcaaattg cagtactctg cgggtgtata cagaatagca
     4621 gaatgggcag acattacgaa tgcacacggt gtggtgggcc caggtattgt tagcggtttg
     4681 aagcaggcgg cagaagaagt aacaaaggaa cctagaggcc ttttgatgtt agcagaattg
     4741 tcatgcaagg gctccctatc tactggagaa tatactaagg gtactgttga cattgcgaag
     4801 agcgacaaag attttgttat cggctttatt gctcaaagag acatgggtgg aagagatgaa
     4861 ggttacgatt ggttgattat gacacccggt gtgggtttag atgacaaggg agacgcattg
     4921 ggtcaacagt atagaaccgt ggatgatgtg gtctctacag gatctgacat tattattgtt
     4981 ggaagaggac tatttgcaaa gggaagggat gctaaggtag agggtgaacg ttacagaaaa
     5041 gcaggctggg aagcatattt gagaagatgc ggccagcaaa actaaaaaac tgtattataa
     5101 gtaaatgcat gtatactaaa ctcacaaatt agagcttcaa tttaattata tcagttatta
     5161 ccctatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggaaatt
     5221 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     5281 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     5341 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     5401 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     5461 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     5521 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     5581 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     5641 gccgcgctta atgcgccgct acagggcgcg tccattcgcc attcaggctg cgcaactgtt
     5701 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     5761 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     5821 cggccagtga attgtaatac gactcactat agggcgaatt ggagctcgcc gggatcgaag
     5881 aaatgatggt aaatgaaata ggaaatcaag gagcatgaag gcaaaagaca aatataaggg
     5941 tcgaacgaaa aataaagtga aaagtgttga tatgatgtat ttggctttgc ggcgccgaaa
     6001 aaacgagttt acgcaattgc acaatcatgc tgactctgtg gcggacccgc gctcttgccg
     6061 gcccggcgat aacgctgggc gtgaggctgt gcccggcgga gttttttgcg cctgcatttt
     6121 ccaaggttta ccctgcgcta aggggcgaga ttggagaagc aataagaatg ccggttgggg
     6181 ttgcgatgat gacgaccacg acaactggtg tcattattta agttgccgaa agaacctgag
     6241 tgcatttgca acatgagtat actagaagaa tgagccaaga cttgcgagac gcgagtttgc
     6301 cggtggtgcg aacaatagag cgaccatgac cttgaaggtg agacgcgcat aaccgctaga
     6361 gtactttgaa gaggaaacag caatagggtt gctaccagta taaatagaca ggtacataca
     6421 acactggaaa tggttgtctg tttgagtacg ctttcaattc atttgggtgt gcactttatt
     6481 atgttacaat atggaaggga actttacact tctcctatgc acatatatta attaaagtcc
     6541 aatgctagta gagaaggggg gtaacacccc tccgcgctct tttccgattt ttttctaaac
     6601 cgtggaatat ttcggatatc cttttgttgt ttccgggtgt acaatatgga cttcctcttt
     6661 tctggcaacc aaacccatac atcgggattc ctataatacc ttcgttggtc tccctaacat
     6721 gtaggtggcg gaggggagat atacaataga acagatacca gacaagacat aatgggctaa
     6781 acaagactac accaattaca ctgcctcatt gatggtggta cataacgaac taatactgta
     6841 gccctagact tgatagccat catcatatcg aagtttcact accctttttc catttgccat
     6901 ctattgaagt aataataggc gcatgcaact tcttttcttt ttttttcttt tctctctccc
     6961 ccgttgttgt ctcaccatat ccgcaatgac aaaaaaatga tggaagacac taaaggaaaa
     7021 aattaacgac aaagacagca ccaacagatg tcgttgttcc agagctgatg aggggtatct
     7081 cgaagcacac gaaacttttt ccttccttca ttcacgcaca ctactctcta atgagcaacg
     7141 gtatacggcc ttccttccag ttacttgaat ttgaaataaa aaaaagtttg ctgtcttgct
     7201 atcaagtata aatagacctg caattattaa tcttttgttt cctcgtcatt gttctcgttc
     7261 cctttcttcc ttgtttcttt ttctgcacaa tatttcaagc tataccaagc atacaatcaa
     7321 ct
//
