LOCUS       sequence-217765-        6759 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cwc25 in pRS315(Leu) vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6759)
  TITLE     Cwc25 plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6759)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pRS315-cwc25 (aa 26-33 deleted)
FEATURES             Location/Qualifiers
     source          1..6759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     98..120
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     112..129
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     113..129
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     136..155
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        136..154
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(1034..1052)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1073..1089)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1073..1089)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1086..1108)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1097..1113
                     /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(1121..1127,1128..1145,1146..1151))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1166..1187
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1304..1321)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1475..2063)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1555..1574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2234..3025,3026..3094))
                     /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     2857..2876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3095..3199)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    3236..3739
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     3781..3799
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3837..3859)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3959..3978
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(4473..5567)
                     /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(5568..5975)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(6276..6731)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6413..6434)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6625..6644
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
       61 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
      121 acggccagtg aattgtaata cgactcacta tagggcgaat tggagctgtt caaggcttac
      181 gccgagtcgc ttactcgatg ctcgctttct ttcttagtgt tgttatccta tataaaacgt
      241 aactgtttag ttatttaagt ccaagtgcat atttacacta taatatataa tataacatct
      301 catctttttt tatttctttt tcttcttctc ccgccgcggg cttacataca gagatataag
      361 aacaagtggt acgtagcatg cagcttctca aaagagatga gcgttcagtt ttgagtacag
      421 ggaagaaagg ccaaatcgag agtacattat catggggtcg ggcgatttga atctgttgaa
      481 atcatggaat cccaagctta tgaaaaatag aaaaaaagta tgggagcagc agaagcttaa
      541 cacgcgattg aaagaaattg aaaaagaacg agaactaaat gaactgctga acgaatcgag
      601 caaggataag ccggagactc tcaagaatga cttggctttg aagaagtcgg gcttggaatg
      661 gatgtatcag gatgcaaaac tgagtgacga aaaggaagac tatttattag gtaagaaaaa
      721 actagattca tctatactaa accaacctgc aacaccgcct gtaagggcgg ccactactat
      781 ttccgcttct ggtgccgcaa catccataag ttcgcagaag aagaagtcaa aattgcttaa
      841 agatgatcca atgagtaagt ttaaggtaac taaacaacaa agaagaactc cagactcaac
      901 aaaaaaaaga gcaatgtcgc aacgagggaa accgttgtcc aagcccgctc cggacctaga
      961 ctactaaact ttccttttct tagagcttgg ttcctcgtcg gatagtggca agtccggtac
     1021 ccagcttttg ttccctttag tgagggttaa tttcgagctt ggcgtaatca tggtcatagc
     1081 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     1141 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     1201 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     1261 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     1321 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     1381 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     1441 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     1501 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     1561 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     1621 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     1681 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     1741 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     1801 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     1861 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     1921 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     1981 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     2041 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     2101 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     2161 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     2221 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     2281 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     2341 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     2401 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     2461 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     2521 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     2581 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     2641 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     2701 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     2761 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     2821 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     2881 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     2941 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     3001 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     3061 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     3121 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     3181 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgggtcc ttttcatcac
     3241 gtgctataaa aataattata atttaaattt tttaatataa atatataaat taaaaataga
     3301 aagtaaaaaa agaaattaaa gaaaaaatag tttttgtttt ccgaagatgt aaaagactct
     3361 agggggatcg ccaacaaata ctacctttta tcttgctctt cctgctctca ggtattaatg
     3421 ccgaattgtt tcatcttgtc tgtgtagaag accacacacg aaaatcctgt gattttacat
     3481 tttacttatc gttaatcgaa tgtatatcta tttaatctgc ttttcttgtc taataaatat
     3541 atatgtaaag tacgcttttt gttgaaattt tttaaacctt tgtttatttt tttttcttca
     3601 ttccgtaact cttctacctt ctttatttac tttctaaaat ccaaatacaa aacataaaaa
     3661 taaataaaca cagagtaaat tcccaaatta ttccatcatt aaaagatacg aggcgcgtgt
     3721 aagttacagg caagcgatcc gtcctaagaa accattatta tcatgacatt aacctataaa
     3781 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
     3841 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
     3901 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
     3961 gcatcagagc agattgtact gagagtgcac catatcgact acgtcgttaa ggccgtttct
     4021 gacagagtaa aattcttgag ggaactttca ccattatggg aaatggttca agaaggtatt
     4081 gacttaaact ccatcaaatg gtcaggtcat tgagtgtttt ttatttgttg tatttttttt
     4141 ttttttagag aaaatcctcc aatatataaa ttaggaatca tagtttcatg attttctgtt
     4201 acacctaact ttttgtgtgg tgccctcctc cttgtcaata ttaatgttaa agtgcaattc
     4261 tttttcctta tcacgttgag ccattagtat caatttgctt acctgtattc ctttacatcc
     4321 tcctttttct ccttcttgat aaatgtatgt agattgcgta tatagtttcg tctaccctat
     4381 gaacatattc cattttgtaa tttcgtgtcg tttctattat gaatttcatt tataaagttt
     4441 atgtacaaat atcataaaaa aagagaatct ttttaagcaa ggattttctt aacttcttcg
     4501 gcgacagcat caccgacttc ggtggtactg ttggaaccac ctaaatcacc agttctgata
     4561 cctgcatcca aaaccttttt aactgcatct tcaatggcct taccttcttc aggcaagttc
     4621 aatgacaatt tcaacatcat tgcagcagac aagatagtgg cgatagggtt gaccttattc
     4681 tttggcaaat ctggagcaga accgtggcat ggttcgtaca aaccaaatgc ggtgttcttg
     4741 tctggcaaag aggccaagga cgcagatggc aacaaaccca aggaacctgg gataacggag
     4801 gcttcatcgg agatgatatc accaaacatg ttgctggtga ttataatacc atttaggtgg
     4861 gttgggttct taactaggat catggcggca gaatcaatca attgatgttg aaccttcaat
     4921 gtagggaatt cgttcttgat ggtttcctcc acagtttttc tccataatct tgaagaggcc
     4981 aaaacattag ctttatccaa ggaccaaata ggcaatggtg gctcatgttg tagggccatg
     5041 aaagcggcca ttcttgtgat tctttgcact tctggaacgg tgtattgttc actatcccaa
     5101 gcgacaccat caccatcgtc ttcctttctc ttaccaaagt aaatacctcc cactaattct
     5161 ctgacaacaa cgaagtcagt acctttagca aattgtggct tgattggaga taagtctaaa
     5221 agagagtcgg atgcaaagtt acatggtctt aagttggcgt acaattgaag ttctttacgg
     5281 atttttagta aaccttgttc aggtctaaca ctaccggtac cccatttagg accacccaca
     5341 gcacctaaca aaacggcatc agccttcttg gaggcttcca gcgcctcatc tggaagtgga
     5401 acacctgtag catcgatagc agcaccacca attaaatgat tttcgaaatc gaacttgaca
     5461 ttggaacgaa catcagaaat agctttaaga accttaatgg cttcggctgt gatttcttga
     5521 ccaacgtggt cacctggcaa aacgacgatc ttcttagggg cagacattag aatggtatat
     5581 ccttgaaata tatatatata tattgctgaa atgtaaaagg taagaaaagt tagaaagtaa
     5641 gacgattgct aaccacctat tggaaaaaac aataggtcct taaataatat tgtcaacttc
     5701 aagtattgtg atgcaagcat ttagtcatga acgcttctct attctatatg aaaagccggt
     5761 tccggcgctc tcacctttcc tttttctccc aatttttcag ttgaaaaagg tatatgcgtc
     5821 aggcgacctc tgaaattaac aaaaaatttc cagtcatcga atttgattct gtgcgatagc
     5881 gcccctgtgt gttctcgtta tgttgaggaa aaaaataatg gttgctaaga gattcgaact
     5941 cttgcatctt acgatacctg agtattccca cagttaactg cggtcaagat atttcttgaa
     6001 tcaggcgcct tagaccgctc ggccaaacaa ccaattactt gttgagaaat agagtataat
     6061 tatcctataa atataacgtt tttgaacaca catgaacaag gaagtacagg acaattgatt
     6121 ttgaagagaa tgtggatttt gatgtaattg ttgggattcc atttttaata aggcaataat
     6181 attaggtatg tagatatact agaagttctc ctcgaccggt cgatatgcgg tgtgaaatac
     6241 cgcacagatg cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt
     6301 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     6361 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     6421 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     6481 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     6541 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     6601 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     6661 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     6721 acagggcgcg tccattcgcc attcaggctg cgcaactgt
//
