LOCUS       sequence-64796-e        7991 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 7991)
  AUTHORS   Finney-Manchester SP, Maheshri N
  TITLE     Harnessing mutagenic homologous recombination for targeted 
            mutagenesis in vivo by TaGTEAM.
  JOURNAL   Nucleic Acids Res. 2013 Mar 7.
  PUBMED    23470991
REFERENCE   2  (bases 1 to 7991)
  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: "Nucleic 
            Acids Res. 2013 Mar 7."
COMMENT     Sequence Label: PRS303-Gal1pr-Mag1-sctetR
FEATURES             Location/Qualifiers
     source          1..7991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    70..573
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        610..714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(715..783,784..1575)
                     /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     complement(933..952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1746..2334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2488..2505
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2658..2663,2664..2681,2682..2688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /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     2701..2723
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2720..2736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2720..2736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2757..2775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3010..3451
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3010..3127
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3422..3445
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             4393..5007
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESG"
     primer_bind     complement(4463..4482)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(5927..5946)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5928..5946)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5953..5970)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5953..5969)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5962..5984)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6114..6569
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6201..6220)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6411..6432
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(6830..7489)
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     promoter        complement(7490..7677)
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     primer_bind     complement(7822..7841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7941..7963
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
      121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
      181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
      241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
      301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
      361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
      421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
      481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
      541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
      601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
      661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
      721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
      781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
      961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1141 ccgaaggagc taaccgcttt ttttcacaac atgggggatc atgtaactcg ccttgatcgt
     1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     1501 ggcagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2101 acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc cacgcttccc
     2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     2221 agggagcttc caggggggaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc cgagcctatg gaaaaacgcc
     2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact
     2641 cattaggcac cccaggcttt acactttatg cttccggctc ctatgttgtg tggaattgtg
     2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcggaatt
     2761 aaccctcact aaagggaaca aaagctgggt accgggcccc ccctcgagca aaaattctta
     2821 cttttttttt ggatggacgc aaagaagttt aataatcata ttacatggca ttaccaccat
     2881 atacatatcc atatctaatc ttacttatat gttgtggaaa tgtaaagagc cccattatct
     2941 tagcctaaaa aaaccttctc tttggaactt tcagtaatac gcttaactgc tcattgctat
     3001 attgaagtac ggattagaag ccgccgagcg ggcgacagcc ctccgacgga agactctcct
     3061 ccgtgcgtcc tcgtcttcac cggtcgcgtt cctgaaacgc agatgtgcct cgcgccgcac
     3121 tgctccgaac aataaagatt ctacaatact agcttttatg gttatgaaga ggaaaaattg
     3181 gcagtaacct ggccccacaa accttcaaat taacgaatca aattaacaac cataggatga
     3241 taatgcgatt agttttttag ccttatttct ggggtaatta atcagcgaag cgatgatttt
     3301 tgatctatta acagatatat aaatggaaaa gctgcataac cactttaact aatactttca
     3361 acattttcag tttgtattac ttcttattca aatgtcataa aagtatcaac aaaaaattgt
     3421 taatatacct ctatacttta acgtcaagga gaaaaaacta taggatccac tagtaacaaa
     3481 atgaaactaa aaagggagta tgatgagtta ataaaagcag acgctgttaa ggaaatagca
     3541 aaagaattag ggtctcgacc tctagaggtt gctcttcctg agaaatatat tgctagacat
     3601 gaagaaaagt tcaatatggc ttgcgaacac attttagaga aagatccatc actttttccc
     3661 atacttaaga ataatgaatt tacgttgtac ttgaaggaga ctcaagtccc taatacactc
     3721 gaagattatt ttattaggct tgcaagcaca attttgtctc aacagatcag tggccaagca
     3781 gctgaaagca tcaaggcaag ggttgtcagt ctttatggcg gtgcatttcc tgattacaaa
     3841 atccttttcg aagacttcaa agacccagca aaatgtgcag aaatcgcaaa atgtggattg
     3901 agtaaaagga aaatgatata tctagagtct cttgctgtct attttactga aaaatataag
     3961 gatatcgaaa agctcttcgg tcaaaaagat aatgatgagg aagtgattga aagtttagtt
     4021 acgaatgtaa aaggtatagg cccatggagt gccaaaatgt tcttgatctc cggattgaaa
     4081 agaatggatg tatttgctcc tgaagatcta ggtattgcta ggggtttttc aaaatacctt
     4141 tcagataagc cagaattgga aaaagaatta atgcgtgaaa gaaaagtagt taaaaagagt
     4201 aagattaagc ataagaaata caactggaaa atatatgacg acgacataat ggaaaaatgc
     4261 tctgaaacat tttctccgta taggtctgtg tttatgttca tactttggag gctcgcgagc
     4321 acaaatacag atgccatgat gaaggcagaa gaaaatttcg tgaaatccgt cgacggtgct
     4381 ggtttaatta actctagatt agataaaagt aaagtgatta acagcgcatt agagctgctt
     4441 aatgaggtcg gaatcgaagg tttaacaacc cgtaaactcg cccagaagct tggtgtagag
     4501 cagcctacat tgtattggca tgtaaaaaat aagcgggctt tgctcgacgc cttagccatt
     4561 gagatgttag ataggcacca tactcacttt tgccctttag aaggggaaag ctggcaagat
     4621 tttttacgta ataacgctaa aagttttaga tgtgctttac taagtcatcg cgatggagca
     4681 aaagtacatt taggtacacg gcctacagaa aaacagtatg aaactctcga aaatcaatta
     4741 gcctttttat gccaacaagg tttttcacta gagaacgcgt tatatgcact cagcgctgtg
     4801 gggcatttta ccttaggttg cgtattggaa gatcaagagc atcaagtcgc taaagaagaa
     4861 agggaaacac ctactactga tagtatgccg ccattattac gacaagctat cgaattattt
     4921 gatcaccaag gtgcagagcc agccttctta ttcggccttg aattgatcat atgcggatta
     4981 gaaaaacaac ttaaatgtga aagtggggac tcaggaggcg gtggatccgg tggaggcggt
     5041 tctggcggtg gaggctctgg aggtggcgga tcgggcggag gtggctctag atctcgtctg
     5101 gacaagagca aagtcataaa ctctgctctg gaattactca atgaagtcgg tatcgaaggc
     5161 ctgacgacaa ggaaactcgc tcaaaagctg ggagttgagc agcctaccct gtactggcac
     5221 gtgaagaaca agcgggccct gctcgatgcc ctggcaatcg agatgctgga caggcatcat
     5281 acccacttct gccccctgga aggcgagtca tggcaagact ttctgcggaa caacgccaag
     5341 tcattccgct gtgctctcct ctcacatcgc gacggggcta aagtgcatct cggcacccgc
     5401 ccaacagaga aacagtacga aaccctggaa aatcagctcg cgttcctgtg tcagcaaggc
     5461 ttctccctgg agaacgcact gtacgctctg tccgccgtgg gccactttac actgggctgc
     5521 gtattggagg atcaggagca tcaagtagca aaagaggaaa gagagacacc taccaccgat
     5581 tctatgcccc cacttctgag acaagcaatt gagctgttcg accatcaggg agccgaacct
     5641 gccttccttt tcggcctgga actaatcata tgtggcctgg agaaacagct aaagtgcgaa
     5701 agcggcgact aataatctgc ttttgtgcgc gtatgtttat gtatgtacct ctctctctat
     5761 ttctattttt aaaccaccct ctcaataaaa taaaaataat aaagtatttt taaggaaaag
     5821 acgtgtttaa gcactgactt tatctacttt ttgtacgttt tcattgatat aatgtgtttt
     5881 gtctctccct tttctacgaa aatttcaaaa attgagctcc aattcgccct atagtgagtc
     5941 gtattacaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     6001 ccaacttaat cgccttgcag cacatccccc cttcgccagc tggcgtaata gcgaagaggc
     6061 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gcgacgcgcc
     6121 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     6181 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     6241 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     6301 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     6361 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     6421 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     6481 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     6541 ttttaacaaa atattaacgt ttacaatttc ctgatgcggt attttctcct tacgcatctg
     6601 tgcggtattt cacaccgcat atgatccgtc gagttcaaga gaaaaaaaaa gaaaaagcaa
     6661 aaagaaaaaa ggaaagcgcg cctcgttcag aatgacacgt atagaatgat gcattacctt
     6721 gtcatcttca gtatcatact gttcgtatac atacttactg acattcatag gtatacatat
     6781 atacacatgt atatatatcg tatgctgcag ctttaaataa tcggtgtcac tacataagaa
     6841 cacctttggt ggagggaaca tcgttggtac cattgggcga ggtggcttct cttatggcaa
     6901 ccgcaagagc cttgaacgca ctctcactac ggtgatgatc attcttgcct cgcagacaat
     6961 caacgtggag ggtaattctg ctagcctctg caaagctttc aagaaaatgc gggatcatct
     7021 cgcaagagag atctcctact ttctcccttt gcaaaccaag ttcgacaact gcgtacggcc
     7081 tgttcgaaag atctaccacc gctctggaaa gtgcctcatc caaaggcgca aatcctgatc
     7141 caaacctttt tactccacgc gccagtaggg cctctttaaa agcttgaccg agagcaatcc
     7201 cgcagtcttc agtggtgtga tggtcgtcta tgtgtaagtc accaatgcac tcaacgatta
     7261 gcgaccagcc ggaatgcttg gccagagcat gtatcatatg gtccagaaac cctatacctg
     7321 tgtggacgtt aatcacttgc gattgtgtgg cctgttctgc tactgcttct gcctcttttt
     7381 ctgggaagat cgagtgctct atcgctaggg gaccaccctt taaagagatc gcaatctgaa
     7441 tcttggtttc atttgtaata cgctttacta gggctttctg ctctgtcatc tttgccttcg
     7501 tttatcttgc ctgctcattt tttagtatat tcttcgaaga aatcacatta ctttatataa
     7561 tgtataattc attatgtgat aatgccaatc gctaagaaaa aaaaagagtc atccgctagg
     7621 tggaaaaaaa aaaatgaaaa tcattaccga ggcataaaaa aatatagagt gtactagagg
     7681 aggccaagag taatagaaaa agaaaattgc gggaaaggac tgtgttatga cttccctgac
     7741 taatgccgtg ttcaaacgat acctggcagt gactcctagc gctcaccaag ctcttaaaac
     7801 ggaattatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     7861 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     7921 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     7981 gaaacgcgcg a
//
