LOCUS       sequence-42848-e        7585 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7585)
  AUTHORS   Larson DR, Zenklusen D, Wu B, Chao JA, Singer RH
  TITLE     Real-time observation of transcription initiation and elongation on 
            an endogenous yeast gene.
  JOURNAL   Science. 2011 Apr 22;332(6028):475-8.
  PUBMED    21512033
REFERENCE   2  (bases 1 to 7585)
  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: "Science."; 
            date: "2011-04-22"; volume: "332(6028)"; pages: "475-8"
COMMENT     Sequence Label: pDZ276 (pURA MET28 PCP-2x-yeGFP)
FEATURES             Location/Qualifiers
     source          1..7585
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             441..1154
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             441..1154
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1193..1209)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1212..1928
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             1935..1952
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1986..2232
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2001..2019)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     2334..2350
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2334..2350
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2384..2400)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(2430..2448)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2469..2485)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2469..2485)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2482..2504)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2493..2509
                     /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(2517..2523,2524..2541,2542..2547))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(2700..2717)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2871..3459)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(3630..4421,4422..4490))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4253..4272
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4491..4595)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    4632..5135
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     5177..5195
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5233..5255)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5355..5374
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5394..5614
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             5615..6418
                     /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(6547..7002)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6684..6705)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6896..6915
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7132..7154
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7146..7163
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7147..7163
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7170..7189
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7170..7188
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        7235..7584
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
ORIGIN
        1 ctagaatgtc caaaaccatc gttctttcgg tcggcgaggc tactcgcact ctgactgaga
       61 tccagtccac cgcagaccgt cagatcttcg aagagaaggt cgggcctctg gtgggtcggc
      121 tgcgcctcac ggcttcgctc cgtcaaaacg gagccaagac cgcgtatcgc gtcaacctaa
      181 aactggatca ggcggacgtc gttgattgct ccaccagcgt ctgcggcgag cttccgaaag
      241 tgcgctacac tcaggtatgg tcgcacgacg tgacaatcgt tgcgaatagc accgaggcct
      301 cgcgcaaatc gttgtacgat ttgaccaagt ccctcgtcgc gacctcgcag gtcgaagatc
      361 ttgtcgtcaa ccttgtgccg ctgggccgtg gatcccccgg gctgcaggaa ttcgatatca
      421 agcttatcga taccgtcgac atgtctaaag gtgaagaatt attcactggt gttgtcccaa
      481 ttttggttga attagatggt gatgttaatg gtcacaaatt ttctgtctcc ggtgaaggtg
      541 aaggtgatgc tacttacggt aaattgacct taaaatttat ttgtactact ggtaaattgc
      601 cagttccatg gccaacctta gtcactactt taacttatgg tgttcaatgt ttttctagat
      661 acccagatca tatgaaacaa catgactttt tcaagtctgc catgccagaa ggttatgttc
      721 aagaaagaac tatttttttc aaagatgacg gtaactacaa gaccagagct gaagtcaagt
      781 ttgaaggtga taccttagtt aatagaatcg aattaaaagg tattgatttt aaagaagatg
      841 gtaacatttt aggtcacaaa ttggaataca actataactc tcacaatgtt tacatcatgg
      901 ctgacaaaca aaagaatggt atcaaagtta acttcaaaat tagacacaac attgaagatg
      961 gttctgttca attagctgac cattatcaac aaaatactcc aattggtgat ggtccagtct
     1021 tgttaccaga caaccattac ttatccactc aatctgcctt atccaaagat ccaaacgaaa
     1081 agagagacca catggtcttg ttagaatttg ttactgctgc tggtattacc catggtatgg
     1141 atgaattgta caaaagatcc cccgggctgc aggaattcga tatcaagctt atcgataccg
     1201 tcgacctcga catgtctaaa ggtgaagaat tattcactgg tgttgtccca attttggttg
     1261 aattagatgg tgatgttaat ggtcacaaat tttctgtctc cggtgaaggt gaaggtgatg
     1321 ctacttacgg taaattgacc ttaaaattta tttgtactac tggtaaattg ccagttccat
     1381 ggccaacctt agtcactact ttcggttatg gtgttcaatg ttttgctaga tacccagatc
     1441 atatgaaaca acatgacttt ttcaagtctg ccatgccaga aggttatgtt caagaaagaa
     1501 ctattttttt caaagatgac ggtaactaca agaccagagc tgaagtcaag tttgaaggtg
     1561 ataccttagt taatagaatc gaattaaaag gtattgattt taaagaagat ggtaacattt
     1621 taggtcacaa attggaatac aactataact ctcacaatgt ttacatcatg gctgacaaac
     1681 aaaagaatgg tatcaaagtt aacttcaaaa ttagacacaa cattgaagat ggttctgttc
     1741 aattagctga ccattatcaa caaaatactc caattggtga tggtccagtc ttgttaccag
     1801 acaaccatta cttatccact caatctgcct tatccaaaga tccaaacgaa aagagagacc
     1861 acatggtctt gttagaattt gttactgctg ctggtattac ccatggtatg gatgaattgt
     1921 acaaataaac cggtcatcat caccatcacc attgagttta aacccgctga tcctagaggg
     1981 ccgcacatgt aattagttat gtcacgctta cattcacgcc ctccccccac atccgctcta
     2041 accgaaaagg aaggagttag acaacctgaa gtctaggtcc ctatttattt ttttatagtt
     2101 atgttagtat taagaacgtt atttatattt caaatttttc ttttttttct gtacagacgc
     2161 gtgtacgcat gtaacattat actgaaaacc ttgcttgaga aggttttggg acgctcgaag
     2221 gctttaattt gcaagctgcg gccctgcatt ggcgcgtgta cgcatgtaac attatactga
     2281 aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcggc cgctctagaa
     2341 ctagtggatc ccccgggctg caggaattcg atatcaagct tatcgatacc gtcgacctcg
     2401 agggggggcc cggtacccag cttttgttcc ctttagtgag ggttaattcc gagcttggcg
     2461 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2521 ataggagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag gtaactcaca
     2581 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2641 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2701 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2761 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2821 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2881 ctcggccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2941 acaggactat aaagatacca ggcgttcccc cctggaagct ccctcgtgcg ctctcctgtt
     3001 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3061 tctcaatgct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3121 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3181 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3241 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3301 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3361 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3421 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3481 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3541 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3601 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3661 tcagcgatct gtctatttcg ttcatccata gttgcctgac tgcccgtcgt gtagataact
     3721 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3781 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3841 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3901 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3961 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4021 acatgatccc ccatgttgtg aaaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4081 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4141 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4201 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4261 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4321 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4381 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4441 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4501 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4561 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4621 gggtcctttt catcacgtgc tataaaaata attataattt aaatttttta atataaatat
     4681 ataaattaaa aatagaaagt aaaaaaagaa attaaagaaa aaatagtttt tgttttccga
     4741 agatgtaaaa gactctaggg ggatcgccaa caaatactac cttttatctt gctcttcctg
     4801 ctctcaggta ttaatgccga attgtttcat cttgtctgtg tagaagacca cacacgaaaa
     4861 tcctgtgatt ttacatttta cttatcgtta atcgaatgta tatctattta atctgctttt
     4921 cttgtctaat aaatatatat gtaaagtacg ctttttgttg aaatttttta aacctttgtt
     4981 tatttttttt tcttcattcc gtaactcttc taccttcttt atttactttc taaaatccaa
     5041 atacaaaaca taaaaataaa taaacacaga gtaaattccc aaattattcc atcattaaaa
     5101 gatacgaggc gcgtgtaagt tacaggcaag cgatccgtcc taagaaacca ttattatcat
     5161 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga
     5221 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     5281 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     5341 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccacg cttttcaatt
     5401 caattcatca tttttttttt attctttttt ttgatttcgg tttctttgaa atttttttga
     5461 ttcggtaatc tccgaacaga aggaagaacg aaggaaggag cacagactta gattggtata
     5521 tatacgcata tgtagtgttg aagaaacatg aaattgccca gtattcttaa cccaactgca
     5581 cagaacaaaa acctgcagga aacgaagata aatcatgtcg aaagctacat ataaggaacg
     5641 tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc acgaaaagca
     5701 aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg agttagttga
     5761 agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga ctgatttttc
     5821 catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt ttttactctt
     5881 cgaagacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact ctgcgggtgt
     5941 atacagaata gcagaatggg cagacattac gaatgcacac ggtgtggtgg gcccaggtat
     6001 tgttagcggt ttgaagcagg cggcagaaga agtaacaaag gaacctagag gccttttgat
     6061 gttagcagaa ttgtcatgca agggctccct atctactgga gaatatacta agggtactgt
     6121 tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa gagacatggg
     6181 tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt tagatgacaa
     6241 gggagacgca ttgggtcaac agtatagaac cgtggatgat gtggtctcta caggatctga
     6301 cattattatt gttggaagag gactatttgc aaagggaagg gatgctaagg tagagggtga
     6361 acgttacaga aaagcaggct gggaagcata tttgagaaga tgcggccagc aaaactaaaa
     6421 aactgtatta taagtaaatg catgtatact aaactcacaa attagagctt caatttaatt
     6481 atatcagtta ttaccctgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     6541 atcaggaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     6601 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     6661 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     6721 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     6781 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     6841 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     6901 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     6961 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcgcgccat tcgccattca
     7021 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
     7081 cgaagggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
     7141 gacgttgtaa aacgacggcc agtgaattgt aatacgactc actatagggc gaattggagc
     7201 tccggatgca agggttcgaa tcccttagct ctcattattt tttgcttttt ctcttgaggt
     7261 cacatgatcg caaaatggca aatggcacgt gaagctgtcg atattgggga actgtggtgg
     7321 ttggcaaatg actaattaag ttagtcaagg cgccatcctc atgaaaactg tgtaacataa
     7381 taaccgaagt gtcgaaaagg tggcaccttg tccaattgaa cacgctcgat gaaaaaaata
     7441 agatatatat aaggttaagt aaagcgtctg ttagaaagga agtttttcct ttttcttgct
     7501 ctcttgtctt ttcatctact atttccttcg tgtaatacag ggtcgtcaga tacatagata
     7561 caattctatt acccccatcc atact
//
