LOCUS       sequence-201325-        8469 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast MCP-GFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8469)
  AUTHORS   Tutucci E, Vera M, Biswas J, Garcia J, Parker R, Singer RH
  TITLE     An improved MS2 system for accurate reporting of the mRNA life 
            cycle.
  JOURNAL   Nat Methods. 2017 Nov 13. doi: 10.1038/nmeth.4502.
  PUBMED    29131164
REFERENCE   2  (bases 1 to 8469)
  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"; journalName: "Nat 
            Methods. 2017 Nov 13. doi: 10.1038/nmeth.4502."
COMMENT     Sequence Label: pET296-YcpLac111 CYC1p-MCP-NLS-2xyeGFP
FEATURES             Location/Qualifiers
     source          1..8469
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    98..119
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(134..139,140..157,158..164)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    172..188
                     /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     177..199
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     196..212
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     196..212
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             903..923
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             930..1640
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIKLKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             1650..2363
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             1650..2360
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     terminator      2394..2640
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2410..2428)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2652..2669)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2652..2668)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2661..2683)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    3425..4587
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN4 centromere fused to the 
                     autonomously replicating sequence ARS1/ARS416"
     rep_origin      3750..4587
                     /label=ARS1
                     /note="S. cerevisiae autonomously replicating sequence 
                     ARS1/ARS416"
     CDS             complement(4949..6043)
                     /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(6044..6449)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     primer_bind     complement(6469..6487)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6555..6659
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6660..6728,6729..7520)
                     /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(6878..6897)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7691..8279
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8180..8199
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8433..8450
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
       61 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
      121 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
      181 gataacaatt tcacacagga aacagctatg accatgatta cgccaagctt gcatgcctgc
      241 agggagcgtt ggttggtgga tcaagcccac gcgtaggcaa tcctcgagca gatccgccgg
      301 gcgtgtatat agcgtggatg gccaggcaac tttagtgctg acacatacag gcatatatat
      361 atgtgtgcga cgacacatga tcatatggca tgcatgtgct ctgtatgtat ataaaactct
      421 tgttttcttc ttttctctaa atattctttc cttatacatt aggtcctttg tagcataaat
      481 tactatactt ctatagacac gcaaacacaa atacacacac taaattaata ggatccatgc
      541 tagccgttaa aatggcttct aactttactc agttcgttct cgtcgacaat ggcggaactg
      601 gcgacgtgac tgtcgcccca agcaacttcg ctaacgggat cgctgaatgg atcagctcta
      661 actcgcgttc acaggcttac aaagtaacct gtagcgttcg tcagagctct gcgcagaatc
      721 gcaaatacac catcaaagtc gaggtgccta aaggcgcctg gcgttcgtac ttaaatatgg
      781 aactaaccat tccaattttc gccacgaatt ccgactgcga gcttattgtt aaggcaatgc
      841 aaggtctcct aaaagatgga aacccgattc cctcagcaat cgcagcaaac tccggcatct
      901 acccaaaaaa aaaaagaaaa gttaccggtt ctaaaggtga agaattattc actggtgttg
      961 tcccaatttt ggttgaatta gatggtgatg ttaatggtca caaattttct gtctccggtg
     1021 aaggtgaagg tgatgctact tacggtaaat tgaccttaaa atttatttgt actactggta
     1081 aattgccagt tccatggcca accttagtca ctactttaac ttatggtgtt caatgttttt
     1141 ctagataccc agatcatatg aaacaacatg actttttcaa gtctgccatg ccagaaggtt
     1201 atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc agagctgaag
     1261 tcaagtttga aggtgatacc ttagttaata gaatcaaatt aaaaggtatt gattttaaag
     1321 aagatggtaa cattttaggt cacaaattgg aatacaacta taactctcac aatgtttaca
     1381 tcatggctga caaacaaaag aatggtatca aagttaactt caaaattaga cacaacattg
     1441 aagatggttc tgttcaatta gctgaccatt atcaacaaaa tactccaatt ggtgatggtc
     1501 cagtcttgtt accagacaac cattacttat ccactcaatc tgccttatcc aaagatccaa
     1561 acgaaaagag agaccacatg gtcttgttag aatttgttac tgctgctggt attacccatg
     1621 gtatggatga attgtacaaa tgtttaaact ctaaaggtga agaattattc actggtgttg
     1681 tcccaatttt ggttgaatta gatggtgatg ttaatggtca caaattttct gtctccggtg
     1741 aaggtgaagg tgatgctact tacggtaaat tgaccttaaa atttatttgt actactggta
     1801 aattgccagt tccatggcca accttagtca ctactttaac ttatggtgtt caatgttttt
     1861 ctagataccc agatcatatg aaacaacatg actttttcaa gtctgccatg ccagaaggtt
     1921 atgttcaaga aagaactatt tttttcaaag atgacggtaa ctacaagacc agagctgaag
     1981 tcaagtttga aggtgatacc ttagttaata gaatcgaatt aaaaggtatt gattttaaag
     2041 aagatggtaa cattttaggt cacaaattgg aatacaacta taactctcac aatgtttaca
     2101 tcatggctga caaacaaaag aatggtatca aagttaactt caaaattaga cacaacattg
     2161 aagatggttc tgttcaatta gctgaccatt atcaacaaaa tactccaatt ggtgatggtc
     2221 cagtcttgtt accagacaac cattacttat ccactcaatc tgccttatcc aaagatccaa
     2281 acgaaaagag agaccacatg gtcttgttag aatttgttac tgctgctggt attacccatg
     2341 gtatggatga attgtacaaa taagtttaaa cccgctgatc ctagagggcc gcatcatgta
     2401 attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga
     2461 aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt
     2521 aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg
     2581 taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg
     2641 agctcgaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     2701 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     2761 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg
     2821 tattttctcc ttacgcatct gtgcggtatt tcacaccgca taatcgctgg gccattctca
     2881 tgaagaatat cttgaattta ttgtcatatt actagttggt gtggaagtcc taatatcggt
     2941 gatcaatata gtggttgaca tgctggctag tcaacattga gccttttgat catgcaaata
     3001 tattacggta ttttacaatc aaatatcaaa cttaactatt gactttataa cttatttagg
     3061 tggtaacatt cttataaaaa agaaaaaaat tactgcaaaa cagtactagc ttttaacttg
     3121 tatcctaggt tatctatgct gtctcaccat agagaatatt acctatttca gaatgtatgt
     3181 ccatgattcg ccgggtaaat acatataata cacaaatctg gcttaataaa gtctataata
     3241 tatctcataa agaagtgcta aattggctag tgctatatat ttttaagaaa atttcttttg
     3301 actaagtcca tatcgacttt gtaaaagttc acattagcat acatatatta cacgagccag
     3361 aaatagtaac ttttgcctaa atcacaaatt gcaaaattta attgcttgca aaaggtcaca
     3421 tgcttataat caactttttt aaaaatttaa aatacttttt tattttttat ttttaaacat
     3481 aaatgaaata atttatttat tgtttatgat taccgaaaca taaaacctgc tcaagaaaaa
     3541 gaaactgttt tgtccttgga aaaaaagcac tacctaggag cggccaaaat gccgaggctt
     3601 tcatagctta aactctttac agaaaatagg cattatagat cagttcgagt tttcttattc
     3661 ttccttccgg ttttatcgtc acagttttac agtaaataag tatcacctct tagagttcga
     3721 tgataagctg tcaaacatga gaattaattc cacatgttaa aatagtgaag gagcatgttc
     3781 ggcacacagt ggaccgaacg tggggtaagt gcactagggt ccggttaaac ggatctcgca
     3841 ttgatgaggc aacgctaatt atcaacatat agattgttat ctatctgcat gaacacgaaa
     3901 tctttacttg acgacttgag gctgatggtg tttatgcaaa gaaaccactg tgtttaatat
     3961 gtgtcactgt ttgatattac tgtcagcgta gaagataata gtaaaagcgg ttaataagtg
     4021 tatttgagat aagtgtgata aagtttttac agcgaaaaga cgataaatac aagaaaatga
     4081 ttacgaggat acggagagag gtatgtacat gtgtatttat atactaagct gccggcggtt
     4141 gtttgcaaga ccgagaaaag gctagcaaga atcgggtcat tgtagcgtat gcgcctgtga
     4201 acattctctt caacaagttt gattccattg cggtgaaatg gtaaaagtca accccctgcg
     4261 atgtatattt tcctgtacaa tcaatcaaaa agccaaatga tttagcatta tctttacatc
     4321 ttgttatttt acagatttta tgtttagatc ttttatgctt gcttttcaaa aggcttgcag
     4381 gcaagtgcac aaacaatact taaataaata ctactcagta ataacctatt tcttagcatt
     4441 tttgacgaaa tttgctattt tgttagagtc ttttacacca tttgtctcca cacctccgct
     4501 tacatcaaca ccaataacgc catttaatct aagcgcatca ccaacatttt ctggcgtcag
     4561 tccaccagct aacataaaat gtaagctctc ggggctctct tgccttccaa cccagtcaga
     4621 aatcgagttc caatccaaaa gttcacctgt cccacctgct tctgaatcaa acaagggaat
     4681 aaacgaatga ggtttctgtg aagctgcact gagtagtatg ttgcagtctt ttggaaatac
     4741 gagtctttta ataactggca aaccgaggaa ctcttggtat tcttgccacg actcatctcc
     4801 atgcagttgg acgatcgatg ataagctgtc aaacatgaga attaattcta ccctatgaac
     4861 atattccatt ttgtaatttc gtgtcgtttc tattatgaat ttcatttata aagtttatgt
     4921 acaaatatca taaaaaaaga gaatcttttt aagcaaggat tttcttaact tcttcggcga
     4981 cagcatcacc gacttcggtg gtactgttgg aaccacctaa atcaccagtt ctgatacctg
     5041 catccaaaac ctttttaact gcatcttcaa tggccttacc ttcttcaggc aagttcaatg
     5101 acaatttcaa catcattgca gcagacaaga tagtggcgat agggttgacc ttattctttg
     5161 gcaaatctgg agcagaaccg tggcatggtt cgtacaaacc aaatgcggtg ttcttgtctg
     5221 gcaaagaggc caaggacgca gatggcaaca aacccaagga acctgggata acggaggctt
     5281 catcggagat gatatcacca aacatgttgc tggtgattat aataccattt aggtgggttg
     5341 ggttcttaac taggatcatg gcggcagaat caatcaattg atgttgaacc ttcaatgtag
     5401 gaaattcgtt cttgatggtt tcctccacag tttttctcca taatcttgaa gaggccaaaa
     5461 cattagcttt atccaaggac caaataggca atggtggctc atgttgtagg gccatgaaag
     5521 cggccattct tgtgattctt tgcacttctg gaacggtgta ttgttcacta tcccaagcga
     5581 caccatcacc atcgtcttcc tttctcttac caaagtaaat acctcccact aattctctga
     5641 caacaacgaa gtcagtacct ttagcaaatt gtggcttgat tggagataag tctaaaagag
     5701 agtcggatgc aaagttacat ggtcttaagt tggcgtacaa ttgaagttct ttacggattt
     5761 ttagtaaacc ttgttcaggt ctaacactac ctgtacccca tttaggacca cccacagcac
     5821 ctaacaaaac ggcatcagcc ttcttggagg cttccagcgc ctcatctgga agtggaacac
     5881 ctgtagcatc gatagcagca ccaccaatta aatgattttc gaaatcgaac ttgacattgg
     5941 aacgaacatc agaaatagct ttaagaacct taatggcttc ggctgtgatt tcttgaccaa
     6001 cgtggtcacc tggcaaaacg acgatcttct taggggcaga cattagaatg gtatatcctt
     6061 gaaatatata tatatattgc tgaaatgtaa aaggtaagaa aagttagaaa gtaagacgat
     6121 tgctaaccac ctattggaaa aaacaatagg tccttaaata atattgtcaa cttcaagtat
     6181 tgtgatgcaa gcatttagtc atgaacgctt ctctattcta tatgaaaagc cggttccggc
     6241 gctctcacct ttcctttttc tcccaatttt tcagttgaaa aaggtatatg cgtcaggcga
     6301 cctctgaaat taacaaaaaa tttccagtca tcgaatttga ttctgtgcga tagcgcccct
     6361 gtgtgttctc gttatgttga ggaaaaaaat aatggttgct aagagattcg aactcttgca
     6421 tcttacgata cctgagtatt cccacagtta attcttgaag acgaaagggc ctcgtgatac
     6481 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     6541 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     6601 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     6661 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     6721 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     6781 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     6841 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     6901 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     6961 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     7021 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     7081 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     7141 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     7201 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     7261 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     7321 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     7381 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     7441 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     7501 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     7561 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     7621 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     7681 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     7741 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     7801 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     7861 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     7921 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     7981 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     8041 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     8101 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     8161 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     8221 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     8281 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     8341 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     8401 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     8461 agcgcccaa
//
