LOCUS       sequence-217759-        6776 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 6776)
  TITLE     Cwc25 plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6776)
  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            "Unpublished"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 26-27 deleted)
FEATURES             Location/Qualifiers
     source          1..6776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      79..534
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(166..185)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     376..397
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        835..1242
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             1243..2337
                     /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"
     primer_bind     complement(2831..2850)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2950..2972
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3010..3028)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3070..3573
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        3610..3714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3715..3783,3784..4575)
                     /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     complement(3933..3952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4746..5334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5235..5254
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5488..5505
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5622..5643
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5658..5663,5664..5681,5682..5688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5696..5712
                     /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     5701..5723
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5720..5736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5720..5736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        5757..5775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6672..6691)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6673..6691)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6698..6715)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6698..6714)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6707..6729)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
       61 agcctgaatg gcgaatggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
      121 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
      181 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
      241 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
      301 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
      361 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
      421 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
      481 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttcctgat
      541 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatcga ccggtcgagg
      601 agaacttcta gtatatctac atacctaata ttattgcctt attaaaaatg gaatcccaac
      661 aattacatca aaatccacat tctcttcaaa atcaattgtc ctgtacttcc ttgttcatgt
      721 gtgttcaaaa acgttatatt tataggataa ttatactcta tttctcaaca agtaattggt
      781 tgtttggccg agcggtctaa ggcgcctgat tcaagaaata tcttgaccgc agttaactgt
      841 gggaatactc aggtatcgta agatgcaaga gttcgaatct cttagcaacc attatttttt
      901 tcctcaacat aacgagaaca cacaggggcg ctatcgcaca gaatcaaatt cgatgactgg
      961 aaattttttg ttaatttcag aggtcgcctg acgcatatac ctttttcaac tgaaaaattg
     1021 ggagaaaaag gaaaggtgag agcgccggaa ccggcttttc atatagaata gagaagcgtt
     1081 catgactaaa tgcttgcatc acaatacttg aagttgacaa tattatttaa ggacctattg
     1141 ttttttccaa taggtggtta gcaatcgtct tactttctaa cttttcttac cttttacatt
     1201 tcagcaatat atatatatat atttcaagga tataccattc taatgtctgc ccctaagaag
     1261 atcgtcgttt tgccaggtga ccacgttggt caagaaatca cagccgaagc cattaaggtt
     1321 cttaaagcta tttctgatgt tcgttccaat gtcaagttcg atttcgaaaa tcatttaatt
     1381 ggtggtgctg ctatcgatgc tacaggtgtt ccacttccag atgaggcgct ggaagcctcc
     1441 aagaaggctg atgccgtttt gttaggtgct gtgggtggtc ctaaatgggg taccggtagt
     1501 gttagacctg aacaaggttt actaaaaatc cgtaaagaac ttcaattgta cgccaactta
     1561 agaccatgta actttgcatc cgactctctt ttagacttat ctccaatcaa gccacaattt
     1621 gctaaaggta ctgacttcgt tgttgtcaga gaattagtgg gaggtattta ctttggtaag
     1681 agaaaggaag acgatggtga tggtgtcgct tgggatagtg aacaatacac cgttccagaa
     1741 gtgcaaagaa tcacaagaat ggccgctttc atggccctac aacatgagcc accattgcct
     1801 atttggtcct tggataaagc taatgttttg gcctcttcaa gattatggag aaaaactgtg
     1861 gaggaaacca tcaagaacga attccctaca ttgaaggttc aacatcaatt gattgattct
     1921 gccgccatga tcctagttaa gaacccaacc cacctaaatg gtattataat caccagcaac
     1981 atgtttggtg atatcatctc cgatgaagcc tccgttatcc caggttcctt gggtttgttg
     2041 ccatctgcgt ccttggcctc tttgccagac aagaacaccg catttggttt gtacgaacca
     2101 tgccacggtt ctgctccaga tttgccaaag aataaggtca accctatcgc cactatcttg
     2161 tctgctgcaa tgatgttgaa attgtcattg aacttgcctg aagaaggtaa ggccattgaa
     2221 gatgcagtta aaaaggtttt ggatgcaggt atcagaactg gtgatttagg tggttccaac
     2281 agtaccaccg aagtcggtga tgctgtcgcc gaagaagtta agaaaatcct tgcttaaaaa
     2341 gattctcttt ttttatgata tttgtacata aactttataa atgaaattca taatagaaac
     2401 gacacgaaat tacaaaatgg aatatgttca tagggtagac gaaactatat acgcaatcta
     2461 catacattta tcaagaagga gaaaaaggag gatgtaaagg aatacaggta agcaaattga
     2521 tactaatggc tcaacgtgat aaggaaaaag aattgcactt taacattaat attgacaagg
     2581 aggagggcac cacacaaaaa gttaggtgta acagaaaatc atgaaactat gattcctaat
     2641 ttatatattg gaggattttc tctaaaaaaa aaaaaataca acaaataaaa aacactcaat
     2701 gacctgacca tttgatggag tttaagtcaa taccttcttg aaccatttcc cataatggtg
     2761 aaagttccct caagaatttt actctgtcag aaacggcctt aacgacgtag tcgatatggt
     2821 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     2881 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     2941 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     3001 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     3061 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
     3121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
     3181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
     3241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
     3301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
     3361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
     3421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
     3481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
     3541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
     3601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     3721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     3961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     4021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     4081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4141 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     4501 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4861 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     5041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     5101 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5221 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     5401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     5461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     5521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     5581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
     5641 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     5701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcgaaatt
     5761 aaccctcact aaagggaaca aaagctgggt accggacttg ccactatccg acgaggaacc
     5821 aagctctaag aaaaggaaag tttagtagtc taggtccgga gcgggcttgg acaacggttt
     5881 ccctcgttgc gacattgctc ttttttttgt tgagtctgga gttcttcttt gttgtttagt
     5941 taccttaaac ttactcattg gatcatcttt aagcaatttt gacttcttct tctgcgaact
     6001 tatggatgtt gcggcaccag aagcggaaat agtagtggcc gcccttacag gcggtgttgc
     6061 aggttggttt agtatagatg aatctagttt tttcttacct aataaatagt cttccttttc
     6121 gtcactcagt tttgcatcct gatacatcca ttccaagccc gacttcttca aagccaagtc
     6181 attcttgaga gtctccggct tatccttgct cgattcgttc agcagttcat ttagttctcg
     6241 ttctttttca atttctttca atcgcgtgtt aagcttctgc tgttcggtaa ttagatcctg
     6301 ctcccatact ttttttctat ttttcataag cttgggattc catgatttca acagattcaa
     6361 atcgcccgac cccatgataa tgtactctcg atttggcctt tcttccctgt actcaaaact
     6421 gaacgctcat ctcttttgag aagctgcatg ctacgtacca cttgttctta tatctctgta
     6481 tgtaagcccg cggcgggaga agaagaaaaa gaaataaaaa aagatgagat gttatattat
     6541 atattatagt gtaaatatgc acttggactt aaataactaa acagttacgt tttatatagg
     6601 ataacaacac taagaaagaa agcgagcagc gagtaagcga ctcggcgtaa gccttgaaca
     6661 gctccaattc gccctatagt gagtcgtatt acaattcact ggccgtcgtt ttacaacgtc
     6721 gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccct
//
