LOCUS       sequence-220924-        8150 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 8150)
  TITLE     Unpublished, Paco
REFERENCE   2  (bases 1 to 8150)
  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"
COMMENT     Sequence Label: pRS415-BUD32
FEATURES             Location/Qualifiers
     source          1..8150
                     /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"
     promoter        complement(1759..2166)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(2467..2922)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2604..2625)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2816..2835
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3048..3070
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3062..3079
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3063..3079
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3089..3108
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3089..3107
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3132..3148
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3133..3149
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5376..5396)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5376..5394)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5415..5431)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5415..5431)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5428..5450)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5439..5455
                     /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(5463..5469,5470..5487,5488..5493))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5508..5529
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5646..5663)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5817..6405)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5897..5916)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6576..7367,7368..7436))
                     /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     7199..7218
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7437..7541)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    7578..8081
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     8123..8141
                     /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 gaatggtata tccttgaaat atatatatat atattgctga
     1801 aatgtaaaag gtaagaaaag ttagaaagta agacgattgc taaccaccta ttggaaaaaa
     1861 caataggtcc ttaaataata ttgtcaactt caagtattgt gatgcaagca tttagtcatg
     1921 aacgcttctc tattctatat gaaaagccgg ttccggcgct ctcacctttc ctttttctcc
     1981 caatttttca gttgaaaaag gtatatgcgt caggcgacct ctgaaattaa caaaaaattt
     2041 ccagtcatcg aatttgattc tgtgcgatag cgcccctgtg tgttctcgtt atgttgagga
     2101 aaaaaataat ggttgctaag agattcgaac tcttgcatct tacgatacct gagtattccc
     2161 acagttaact gcggtcaaga tatttcttga atcaggcgcc ttagaccgct cggccaaaca
     2221 accaattact tgttgagaaa tagagtataa ttatcctata aatataacgt ttttgaacac
     2281 acatgaacaa ggaagtacag gacaattgat tttgaagaga atgtggattt tgatgtaatt
     2341 gttgggattc catttttaat aaggcaataa tattaggtat gtagatatac tagaagttct
     2401 cctcgaccgg tcgatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     2461 atcaggaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     2521 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     2581 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     2641 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     2701 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     2761 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     2821 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2881 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtccattcgc cattcaggct
     2941 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     3001 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     3061 ttgtaaaacg acggccagtg agcgcgcgta atacgactca ctatagggcg aattgggtac
     3121 cgggcccccc ctcgaggtcg acggtatcga taagcttgat atcgaattcc tgcagcccgg
     3181 gggatcagtt ttagtattct atcatggggg tggttatgct cttaaattga cgcctacgtc
     3241 gttttctttc ttgaataata tgcgaaatgc ctttcctaaa atggccattc ttgtaccaga
     3301 ttataccgtt acagctaccg atgaccagtc gaaaaagtac cctctgcaaa ttttacaaaa
     3361 tgtggccatt tttgattacg ttgttaagac tatggggtgc aaaaatgtag tgattatggg
     3421 ggattctgct ggtggcaatg ctgttttgaa cattgtttta tatttaagaa agtgtcacag
     3481 agaaatatac ccgaaaaaag tcatagccat aagtccctgg gccaatgcca ctttcttcca
     3541 cgagggagaa aaggaatata tgcaaggaac acaagaatgg gatggtctat gtttgaagag
     3601 tcattcgatg tttgggcgaa tgtttgtagg aaacaaccca aacgtcgatt ttaccagcga
     3661 cccttttgtt aatattgaaa agaatttcga aactaaaatg tggcaagata ttctgaaaaa
     3721 gtgctctgtg atgattactt atggtagtga tgagttgttg agctttcaaa acaagatctt
     3781 ggcaaagaaa atgagtgacg ctagcgaggg atgtaaccac tttaccgcca aaaatgtgct
     3841 ggttgaacac caaggttatc atacgggccc aatattaaac tactctcgta atatggatcg
     3901 ctggaccaac ataccatcta tcgctcgcat tcttgagttt atgcaatcct gaatttgtca
     3961 taatgacaat tacataaccc cttttatata cataaatatt tgggtcagca caaatttatt
     4021 ttgattttcc attattttag cagcaacaag gttgaaaaaa tcgccggcgc atcaccaact
     4081 tgaaggacaa agataacctt acttgaaatc aaataagtga tgaatacgac agcggaaaag
     4141 gttatacgct ttgctcatta tgacgcaaga attcattgat aaagtatctt cctacctgac
     4201 tcctgatgtg gacattgcac ctatttctca aggtgcagaa gccattgttt tcacaacgac
     4261 tactcaccca tatctcccaa gagcaaagga ttctcatcaa aagtatatta tcaaatatag
     4321 gccaccaaag cgttatagac atccacaaat agaccaggca ctaacaaaac atcgtacgtt
     4381 gaatgagtcg cgtttattgg ccaaattata cttgattcca gggctatgtg tccctcaact
     4441 gatagcgtgt gatccataca atggattcat ttggttagag ttccttggag aagatcttcc
     4501 cggagggcac ggttttagta acctaaagaa cttcctttgg atgcatgacc aagatccata
     4561 tagtgatctt gtagcaacta cactacggaa agtggggcgc caaattgggt tgttgcactg
     4621 gaatgactac tgtcatggtg atttgacaag ttctaacatt gttctcgtgc gagatggtgc
     4681 gagatggacg cctcatttga ttgatttcgg tctgggctca gtttcaaacc tggtcgaaga
     4741 taaaggcgtc gatttatacg tcttagagag agctatttta agtacacatt cgaagcatgc
     4801 ggaaaaatac aatgcttgga tcatggaggg gttcgaagag gtctatcgtg aacaaggtgc
     4861 gaaaggtgcc aagaaactga aagaagttac caaaagattc gaagaggtca ggttgcgtgg
     4921 tcgtaagaga agtatgctag gataatataa atgctagcgt actgcctgta tatcgctgca
     4981 cataatatgt atctataatt tatattaacg tacataactt ttaggtatgt taccatcaaa
     5041 gcgtgtccga gttattgaga ttttgacaaa ttcagacaaa tcatttttga agaaccatat
     5101 atataaatat ataaagaaaa gaaaggtaaa ccaaagacag caaacaatcg aaaagtggac
     5161 aaccagcaat ggtagattca attcaccgta ttgcatcggc gttggatacg gccaaagtaa
     5221 taacgaggga agctgccgcg gtagccacct ctaagctggg tgaatcctcc tatacttatt
     5281 attctcaaaa catcaatcct caacagttag tcaccttgtt aaattctagg aactctagag
     5341 cggccgccac cgcggtggag ctccagcttt tgttcccttt agtgagggtt aattgcgcgc
     5401 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     5461 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     5521 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5581 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     5641 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     5701 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     5761 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     5821 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     5881 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     5941 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     6001 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     6061 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     6121 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     6181 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     6241 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     6301 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     6361 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6421 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6481 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     6541 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6601 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     6661 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     6721 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     6781 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     6841 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     6901 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     6961 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     7021 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     7081 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     7141 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     7201 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     7261 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     7321 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     7381 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7441 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7501 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7561 ccacctgggt ccttttcatc acgtgctata aaaataatta taatttaaat tttttaatat
     7621 aaatatataa attaaaaata gaaagtaaaa aaagaaatta aagaaaaaat agtttttgtt
     7681 ttccgaagat gtaaaagact ctagggggat cgccaacaaa tactaccttt tatcttgctc
     7741 ttcctgctct caggtattaa tgccgaattg tttcatcttg tctgtgtaga agaccacaca
     7801 cgaaaatcct gtgattttac attttactta tcgttaatcg aatgtatatc tatttaatct
     7861 gcttttcttg tctaataaat atatatgtaa agtacgcttt ttgttgaaat tttttaaacc
     7921 tttgtttatt tttttttctt cattccgtaa ctcttctacc ttctttattt actttctaaa
     7981 atccaaatac aaaacataaa aataaataaa cacagagtaa attcccaaat tattccatca
     8041 ttaaaagata cgaggcgcgt gtaagttaca ggcaagcgat ccgtcctaag aaaccattat
     8101 tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
//
