LOCUS       sequence-267914-        7346 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7346)
  AUTHORS   Ro DK, Paradise EM, Ouellet M, Fisher KJ, Newman KL, Ndungu JM, Ho 
            KA, Eachus RA, Ham TS, Kirby J, Chang MC, Withers ST, Shiba Y, 
            Sarpong R, Keasling JD
  TITLE     Production of the antimalarial drug precursor artemisinic acid in 
            engineered yeast.
  JOURNAL   Nature. 2006 Apr 13. 440(7086):940-3.
  PUBMED    16612385
REFERENCE   2  (bases 1 to 7346)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2006 Apr 13. 440(7086):940-3."
COMMENT     Sequence Label: pESC-leu2d(empty)
FEATURES             Location/Qualifiers
     source          1..7346
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(664..1758)
                     /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"
     rep_origin      complement(2066..2521)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2203..2224)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2415..2434
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(2600..2765)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2686..2710
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2913..2936)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        2983..3647
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    3199..3316
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3611..3634
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     3658..3677
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3658..3676
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3694..3723
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      3753..3942
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4014..4031)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4185..4773)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4265..4284)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4944..5735,5736..5804))
                     /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     5567..5586
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5805..5909)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5936..7278
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(6899..6946)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     7319..7337
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatcga ctacgtcgtt aaggccgttt ctgacagagt aaaattcttg agggaacttt
      241 caccattatg ggaaatggtt caagaaggta ttgacttaaa ctccatcaaa tggtcaggtc
      301 attgagtgtt ttttatttgt tgtatttttt tttttttaga gaaaatcctc caatatataa
      361 attaggaatc atagtttcat gattttctgt tacacctaac tttttgtgtg gtgccctcct
      421 ccttgtcaat attaatgtta aagtgcaatt ctttttcctt atcacgttga gccattagta
      481 tcaatttgct tacctgtatt cctttacatc ctcctttttc tccttcttga taaatgtatg
      541 tagattgcgt atatagtttc gtctacccta tgaacatatt ccattttgta atttcgtgtc
      601 gtttctatta tgaatttcat ttataaagtt tatgtacaaa tatcataaaa aaagagaatc
      661 tttttaagca aggattttct taacttcttc ggcgacagca tcaccgactt cggtggtact
      721 gttggaacca cctaaatcac cagttctgat acctgcatcc aaaacctttt taactgcatc
      781 ttcaatggcc ttaccttctt caggcaagtt caatgacaat ttcaacatca ttgcagcaga
      841 caagatagtg gcgatagggt tgaccttatt ctttggcaaa tctggagcag aaccgtggca
      901 tggttcgtac aaaccaaatg cggtgttctt gtctggcaaa gaggccaagg acgcagatgg
      961 caacaaaccc aaggaacctg ggataacgga ggcttcatcg gagatgatat caccaaacat
     1021 gttgctggtg attataatac catttaggtg ggttgggttc ttaactagga tcatggcggc
     1081 agaatcaatc aattgatgtt gaaccttcaa tgtagggaat tcgttcttga tggtttcctc
     1141 cacagttttt ctccataatc ttgaagaggc caaaacatta gctttatcca aggaccaaat
     1201 aggcaatggt ggctcatgtt gtagggccat gaaagcggcc attcttgtga ttctttgcac
     1261 ttctggaacg gtgtattgtt cactatccca agcgacacca tcaccatcgt cttcctttct
     1321 cttaccaaag taaatacctc ccactaattc tctgacaaca acgaagtcag tacctttagc
     1381 aaattgtggc ttgattggag ataagtctaa aagagagtcg gatgcaaagt tacatggtct
     1441 taagttggcg tacaattgaa gttctttacg gatttttagt aaaccttgtt caggtctaac
     1501 actaccggta ccccatttag gaccacccac agcacctaac aaaacggcat cagccttctt
     1561 ggaggcttcc agcgcctcat ctggaagtgg aacacctgta gcatcgatag cagcaccacc
     1621 aattaaatga ttttcgaaat cgaacttgac attggaacga acatcagaaa tagctttaag
     1681 aaccttaatg gcttcggctg tgatttcttg accaacgtgg tcacctggca aaacgacgat
     1741 cttcttaggg gcagacatta caatggtata tccttgaaat atatataaaa aaaggcgcct
     1801 tagaccgctc ggccaaacaa ccaattactt gttgagaaat agagtataat tatcctataa
     1861 atataacgtt tttgaacaca catgaacaag gaagtacagg acaattgatt ttgaagagaa
     1921 tgtggatttt gatgtaattg ttgggattcc atttttaata aggcaataat attaggtatg
     1981 tagatatact agaagttctc ctcgaccggt cgatatgcgg tgtgaaatac cgcacagatg
     2041 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt aaaattcgcg
     2101 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     2161 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     2221 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     2281 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     2341 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     2401 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     2461 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     2521 tccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
     2581 tattacgcca gctgaattgg agcgacctca tgctatacct gagaaagcaa cctgacctac
     2641 aggaaagagt tactcaagaa taagaatttt cgttttaaaa cctaagagtc actttaaaat
     2701 ttgtatacac ttattttttt tataacttat ttaataataa aaatcataaa tcataagaaa
     2761 ttcgcttatt tagaagtgtc aacaacgtat ctaccaacga tttgaccctt ttccatcttt
     2821 tcgtaaattt ctggcaaggt agacaagccg acaaccttga ttggagactt gaccaaacct
     2881 ctggcgaaga attgttaatt aagagctcag atcttatcgt cgtcatcctt gtaatccatc
     2941 gatactagtg cggccgccct ttagtgaggg ttgaattcga attttcaaaa attcttactt
     3001 tttttttgga tggacgcaaa gaagtttaat aatcatatta catggcatta ccaccatata
     3061 catatccata tacatatcca tatctaatct tacttatatg ttgtggaaat gtaaagagcc
     3121 ccattatctt agcctaaaaa aaccttctct ttggaacttt cagtaatacg cttaactgct
     3181 cattgctata ttgaagtacg gattagaagc cgccgagcgg gtgacagccc tccgaaggaa
     3241 gactctcctc cgtgcgtcct cgtcttcacc ggtcgcgttc ctgaaacgca gatgtgcctc
     3301 gcgccgcact gctccgaaca ataaagattc tacaatacta gcttttatgg ttatgaagag
     3361 gaaaaattgg cagtaacctg gccccacaaa ccttcaaatg aacgaatcaa attaacaacc
     3421 ataggatgat aatgcgatta gttttttagc cttatttctg gggtaattaa tcagcgaagc
     3481 gatgattttt gatctattaa cagatatata aatgcaaaaa ctgcataacc actttaacta
     3541 atactttcaa cattttcggt ttgtattact tcttattcaa atgtaataaa agtatcaaca
     3601 aaaaattgtt aatatacctc tatactttaa cgtcaaggag aaaaaacccc ggatccgtaa
     3661 tacgactcac tatagggccc gggcgtcgac atggaacaga agttgatttc cgaagaagac
     3721 ctcgagtaag cttggtaccg cggctagcta agatccgctc taaccgaaaa ggaaggagtt
     3781 agacaacctg aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg
     3841 ttatttatat ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc atgtaacatt
     3901 atactgaaaa ccttgcttga gaaggttttg ggacgctcga agatccagct gcattaatga
     3961 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4021 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4081 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4141 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4201 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4261 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4321 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4381 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4441 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4501 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4561 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4621 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4681 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4741 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4801 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4861 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4921 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4981 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5041 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5101 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5161 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5221 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5281 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5341 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5401 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5461 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5521 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5581 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5641 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5701 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5761 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5821 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5881 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgaacga
     5941 agcatctgtg cttcattttg tagaacaaaa atgcaacgcg agagcgctaa tttttcaaac
     6001 aaagaatctg agctgcattt ttacagaaca gaaatgcaac gcgaaagcgc tattttacca
     6061 acgaagaatc tgtgcttcat ttttgtaaaa caaaaatgca acgcgagagc gctaattttt
     6121 caaacaaaga atctgagctg catttttaca gaacagaaat gcaacgcgag agcgctattt
     6181 taccaacaaa gaatctatac ttcttttttg ttctacaaaa atgcatcccg agagcgctat
     6241 ttttctaaca aagcatctta gattactttt tttctccttt gtgcgctcta taatgcagtc
     6301 tcttgataac tttttgcact gtaggtccgt taaggttaga agaaggctac tttggtgtct
     6361 attttctctt ccataaaaaa agcctgactc cacttcccgc gtttactgat tactagcgaa
     6421 gctgcgggtg cattttttca agataaaggc atccccgatt atattctata ccgatgtgga
     6481 ttgcgcatac tttgtgaaca gaaagtgata gcgttgatga ttcttcattg gtcagaaaat
     6541 tatgaacggt ttcttctatt ttgtctctat atactacgta taggaaatgt ttacattttc
     6601 gtattgtttt cgattcactc tatgaatagt tcttactaca atttttttgt ctaaagagta
     6661 atactagaga taaacataaa aaatgtagag gtcgagttta gatgcaagtt caaggagcga
     6721 aaggtggatg ggtaggttat atagggatat agcacagaga tatatagcaa agagatactt
     6781 ttgagcaatg tttgtggaag cggtattcgc aatattttag tagctcgtta cagtccggtg
     6841 cgtttttggt tttttgaaag tgcgtcttca gagcgctttt ggttttcaaa agcgctctga
     6901 agttcctata ctttctagag aataggaact tcggaatagg aacttcaaag cgtttccgaa
     6961 aacgagcgct tccgaaaatg caacgcgagc tgcgcacata cagctcactg ttcacgtcgc
     7021 acctatatct gcgtgttgcc tgtatatata tatacatgag aagaacggca tagtgcgtgt
     7081 ttatgcttaa atgcgtactt atatgcgtct atttatgtag gatgaaaggt agtctagtac
     7141 ctcctgtgat attatcccat tccatgcggg gtatcgtatg cttccttcag cactaccctt
     7201 tagctgttct atatgctgcc actcctcaat tggattagtc tcatccttca atgctatcat
     7261 ttcctttgat attggatcat actaagaaac cattattatc atgacattaa cctataaaaa
     7321 taggcgtatc acgaggccct ttcgtc
//
