LOCUS       sequence-217763-        6770 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 6770)
  TITLE     Cwc25 plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6770)
  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            "Unpublished"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 26-29 deleted)
FEATURES             Location/Qualifiers
     source          1..6770
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     62..84
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     76..93
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     77..93
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     100..119
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        100..118
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(1010..1028)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1049..1065)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1049..1065)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1062..1084)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1073..1089
                     /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(1097..1103,1104..1121,1122..1127))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1142..1163
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1280..1297)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1451..2039)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1531..1550)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2210..3001,3002..3070))
                     /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     2833..2852
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3071..3175)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    3212..3715
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     3757..3775
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3813..3835)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3935..3954
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(4448..5542)
                     /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(5543..5950)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(6251..6706)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6388..6409)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6600..6619
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
       61 tcccagtcac gacgttgtaa aacgacggcc agtgaattgt aatacgactc actatagggc
      121 gaattggagc tgttcaaggc ttacgccgag tcgcttactc gctgctcgct ttctttctta
      181 gtgttgttat cctatataaa acgtaactgt ttagttattt aagtccaagt gcatatttac
      241 actataatat ataatataac atctcatctt tttttatttc tttttcttct tctcccgccg
      301 cgggcttaca tacagagata taagaacaag tggtacgtag catgcagctt ctcaaaagag
      361 atgagcgttc agttttgagt acagggaaga aaggccaaat cgagagtaca ttatcatggg
      421 gtcgggcgat ttgaatctgt tgaaatcatg gaatcccaag cttatgaaaa atagaaaaaa
      481 agtatgggag ctaattaccg aacagcagaa gcttaacacg cgattgaaag aaattgaaaa
      541 agaacgagaa ctaaatgaac tgctgaacga atcgagcaag gataagccgg agactctcaa
      601 gaatgacttg gctttgaaga agtcgggctt ggaatggatg tatcaggatg caaaactgag
      661 tgacgaaaag gaagactatt tattaggtaa gaaaaaacta gattcatcta tactaaacca
      721 acctgcaaca ccgcctgtaa gggcggccac tactatttcc gcttctggtg ccgcaacatc
      781 cataagttcg cagaagaaga agtcaaaatt gcttaaagat gatccaatga gtaagtttaa
      841 ggtaactaaa caacaaagaa gaactccaga ctcaacaaaa aaaagagcaa tgtcgcaacg
      901 agggaaaccg ttgtccaagc ccgctccgga cctagactac taaactttcc ttttcttaga
      961 gcttggttcc tcgtcggata gtggcaagtc cggtacccag cttttgttcc ctttagtgag
     1021 ggttaatttc gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     1081 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     1141 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     1201 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     1261 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1321 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1381 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1441 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1501 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1561 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     1621 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     1681 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     1741 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     1801 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     1861 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     1921 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     1981 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     2041 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     2101 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     2161 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2221 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2281 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2341 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2401 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2461 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     2521 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     2581 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     2641 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     2701 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     2761 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     2821 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     2881 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     2941 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3001 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     3061 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     3121 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     3181 ttccccgaaa agtgccacct gggtcctttt catcacgtgc tataaaaata attataattt
     3241 aaatttttta atataaatat ataaattaaa aatagaaagt aaaaaaagaa attaaagaaa
     3301 aaatagtttt tgttttccga agatgtaaaa gactctaggg ggatcgccaa caaatactac
     3361 cttttatctt gctcttcctg ctctcaggta ttaatgccga attgtttcat cttgtctgtg
     3421 tagaagacca cacacgaaaa tcctgtgatt ttacatttta cttatcgtta atcgaatgta
     3481 tatctattta atctgctttt cttgtctaat aaatatatat gtaaagtacg ctttttgttg
     3541 aaatttttta aacctttgtt tatttttttt tcttcattcc gtaactcttc taccttcttt
     3601 atttactttc taaaatccaa atacaaaaca taaaaataaa taaacacaga gtaaattccc
     3661 aaattattcc atcattaaaa gatacgaggc gcgtgtaagt tacaggcaag cgatccgtcc
     3721 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     3781 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
     3841 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
     3901 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
     3961 gtgcaccata tcgactacgt cgttaaggcc gtttctgaca gagtaaaatt cttgagggaa
     4021 ctttcaccat tatgggaaat ggttcaagaa ggtattgact taaactccat caaatggtca
     4081 ggtcattgag tgttttttat ttgttgtatt tttttttttt tagagaaaat cctccaatat
     4141 ataaattagg aatcatagtt tcatgatttt ctgttacacc taactttttg tgtggtgccc
     4201 tcctccttgt caatattaat gttaaagtgc aattcttttt ccttatcacg ttgagccatt
     4261 agtatcaatt tgcttacctg tattccttta catcctcctt tttctccttc ttgataaatg
     4321 tatgtagatt gcgtatatag tttcgtctac cctatgaaca tattccattt tgtaatttcg
     4381 tgtcgtttct attatgaatt tcatttataa agtttatgta caaatatcat aaaaaaagag
     4441 aatcttttta agcaaggatt ttcttaactt cttcggcgac agcatcaccg acttcggtgg
     4501 tactgttgga accacctaaa tcaccagttc tgatacctgc atccaaaacc tttttaactg
     4561 catcttcaat ggccttacct tcttcaggca agttcaatga caatttcaac atcattgcag
     4621 cagacaagat agtggcgata gggttgacct tattctttgg caaatctgga gcagaaccgt
     4681 ggcatggttc gtacaaacca aatgcggtgt tcttgtctgg caaagaggcc aaggacgcag
     4741 atggcaacaa acccaaggaa cctgggataa cggaggcttc atcggagatg atatcaccaa
     4801 acatgttgct ggtgattata ataccattta ggtgggttgg gttcttaact aggatcatgg
     4861 cggcagaatc aatcaattga tgttgaacct tcaatgtagg gaattcgttc ttgatggttt
     4921 cctccacagt ttttctccat aatcttgaag aggccaaaac attagcttta tccaaggacc
     4981 aaataggcaa tggtggctca tgttgtaggg ccatgaaagc ggccattctt gtgattcttt
     5041 gcacttctgg aacggtgtat tgttcactat cccaagcgac accatcacca tcgtcttcct
     5101 ttctcttacc aaagtaaata cctcccacta attctctgac aacaacgaag tcagtacctt
     5161 tagcaaattg tggcttgatt ggagataagt ctaaaagaga gtcggatgca aagttacatg
     5221 gtcttaagtt ggcgtacaat tgaagttctt tacggatttt tagtaaacct tgttcaggtc
     5281 taacactacc ggtaccccat ttaggaccac ccacagcacc taacaaaacg gcatcagcct
     5341 tcttggaggc ttccagcgcc tcatctggaa gtggaacacc tgtagcatcg atagcagcac
     5401 caccaattaa atgattttcg aaatcgaact tgacattgga acgaacatca gaaatagctt
     5461 taagaacctt aatggcttcg gctgtgattt cttgaccaac gtggtcacct ggcaaaacga
     5521 cgatcttctt aggggcagac attagaatgg tatatccttg aaatatatat atatatattg
     5581 ctgaaatgta aaaggtaaga aaagttagaa agtaagacga ttgctaacca cctattggaa
     5641 aaaacaatag gtccttaaat aatattgtca acttcaagta ttgtgatgca agcatttagt
     5701 catgaacgct tctctattct atatgaaaag ccggttccgg cgctctcacc tttccttttt
     5761 ctcccaattt ttcagttgaa aaaggtatat gcgtcaggcg acctctgaaa ttaacaaaaa
     5821 atttccagtc atcgaatttg attctgtgcg atagcgcccc tgtgtgttct cgttatgttg
     5881 aggaaaaaaa taatggttgc taagagattc gaactcttgc atcttacgat acctgagtat
     5941 tcccacagtt aactgcggtc aagatatttc ttgaatcagg cgccttagac cgctcggcca
     6001 aacaaccaat tacttgttga gaaatagagt ataattatcc tataaatata acgtttttga
     6061 acacacatga acaaggaagt acaggacaat tgattttgaa gagaatgtgg attttgatgt
     6121 aattgttggg attccatttt taataaggca ataatattag gtatgtagat atactagaag
     6181 ttctcctcga ccggtcgata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
     6241 ccgcatcagg aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     6301 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     6361 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     6421 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     6481 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     6541 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     6601 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     6661 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccat tcgccattca
     6721 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
//
