LOCUS       sequence-160276-        6893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Destination vector for integration at yeast URA3 locus, expression 
            from the GPD promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6893)
  TITLE     Nemhauser plasmids
REFERENCE   2  (bases 1 to 6893)
  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: pGP6G-ccdB
FEATURES             Location/Qualifiers
     source          1..6893
                     /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"
     promoter        226..442
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             443..1246
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLATGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     terminator      1247..1434
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1302..1326)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        1459..2102
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     1996..2019
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     2108..2124
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2108..2124
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2132..2256
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2281..2286,2287..2304,2305..2311)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2365..3024
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2431..2450)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3369..3674
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3553..3572
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3715..3839)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3872..3888)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3873..3889)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      3891..4137
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3907..3925)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4672..4688)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4672..4688)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4685..4707)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4696..4712
                     /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(4720..4726,4727..4744,4745..4750))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4903..4920)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5074..5662)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5833..6624,6625..6693))
                     /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     6456..6475
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6694..6798)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6866..6884
                     /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 accatagttt aaactgtggc tgtggtttca gggtccataa agcttttcaa ttcatctttt
      241 ttttttttgt tctttttttt gattccggtt tctttgaaat ttttttgatt cggtaatctc
      301 cgagcagaag gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg
      361 tggtgttgaa gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac
      421 ctgcaggaaa cgaagataaa tcatgtcgaa agctacatat aaggaacgtg ctgctactca
      481 tcctagtcct gttgctgcca agctatttaa tatcatgcac gaaaagcaaa caaacttgtg
      541 tgcttcattg gatgttcgta ccaccaagga attactggag ttagttgaag cattaggtcc
      601 caaaatttgt ttactaaaaa cacatgtgga tatcttgact gatttttcca tggagggcac
      661 agttaagccg ctaaaggcat tatccgccaa gtacaatttt ttactcttcg aagacagaaa
      721 atttgctgac attggtaata cagtcaaatt gcagtactct gcgggtgtat acagaatagc
      781 agaatgggca gacattacga atgcacacgg tgtggtgggc ccaggtattg ttagcggttt
      841 gaagcaggcg gcggaagaag taacaaagga acctagaggc cttttgatgt tagcagaatt
      901 gtcatgcaag ggctccctag ctactggaga atatactaag ggtactgttg acattgcgaa
      961 gagcgacaaa gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga
     1021 aggttacgat tggttgatta tgacacccgg tgtgggttta gatgacaagg gagacgcatt
     1081 gggtcaacag tatagaaccg tggatgatgt ggtctctaca ggatctgaca ttattattgt
     1141 tggaagagga ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa
     1201 agcaggctgg gaagcatatt tgagaagatg cggccagcaa aactaagcga atttcttatg
     1261 atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca aattttaaag
     1321 tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc tgtaggtcag
     1381 gttgctttct caggtatagc atgaggtcgc tcttattgac cacacctccg ccagggtttt
     1441 cccagtcacg acagtttatc attatcaata ctcgccattt caaagaatac gtaaataatt
     1501 aatagtagtg attttcctaa ctttatttag tcaaaaaatt agccttttaa ttctgctgta
     1561 acccgtacat gcccaaaata gggggcgggt tacacagaat atataacatc gtaggtgtct
     1621 gggtgaacag tttattcctg gcatccacta aatataatgg agcccgcttt ttaagctggc
     1681 atccagaaaa aaaaagaatc ccagcaccaa aatattgttt tcttcaccaa ccatcagttc
     1741 ataggtccat tctcttagcg caactacaga gaacaggggc acaaacaggc aaaaaacggg
     1801 cacaacctca atggagtgat gcaacctgcc tggagtaaat gatgacacaa ggcaattgac
     1861 ccacgcatgt atctatctca ttttcttaca ccttctatta ccttctgctc tctctgattt
     1921 ggaaaaagct gaaaaaaaag gttgaaacca gttccctgaa attattcccc tacttgacta
     1981 ataagtatat aaagacggta ggtattgatt gtaattctgt aaatctattt cttaaacttc
     2041 ttaaattcta cttttatagt tagtcttttt tttagtttta aaacaccaga acttagtttc
     2101 gacggattct agaactagtg gatcccccat cacaagtttg tacaaaaaag ctgaacgaga
     2161 aacgtaaaat gatataaata tcaatatatt aaattagatt ttgcataaaa aacagactac
     2221 ataatactgt aaaacacaac atatccagtc actatggcgg ccgcattagg caccccaggc
     2281 tttacacttt atgcttccgg ctcgtataat gtgtggattt tgagttagga tccgtcgaga
     2341 ttttcaggag ctaaggaagc taaaatggag aaaaaaatca ctggatatac caccgttgat
     2401 atatcccaat ggcatcgtaa agaacatttt gaggcatttc agtcagttgc tcaatgtacc
     2461 tataaccaga ccgttcagct ggatattacg gcctttttaa agaccgtaaa gaaaaataag
     2521 cacaagtttt atccggcctt tattcacatt cttgcccgcc tgatgaatgc tcatccggaa
     2581 ttccgtatgg caatgaaaga cggtgagctg gtgatatggg atagtgttca cccttgttac
     2641 accgttttcc atgagcaaac tgaaacgttt tcatcgctct ggagtgaata ccacgacgat
     2701 ttccggcagt ttctacacat atattcgcaa gatgtggcgt gttacggtga aaacctggcc
     2761 tatttcccta aagggtttat tgagaatatg tttttcgtct cagccaatcc ctgggtgagt
     2821 ttcaccagtt ttgatttaaa cgtggccaat atggacaact tcttcgcccc cgttttcacc
     2881 atgggcaaat attatacgca aggcgacaag gtgctgatgc cgctggcgat tcaggttcat
     2941 catgccgtct gtgatggctt ccatgtcggc agaatgctta atgaattaca acagtactgc
     3001 gatgagtggc agggcggggc gtaaacgccg cgtggatccg gcttactaaa agccagataa
     3061 cagtatgcgt atttgcgcgc tgatttttgc ggtataagaa tatatactga tatgtatacc
     3121 cgaagtatgt caaaaagagg tatgctatga agcagcgtat tacagtgaca gttgacagcg
     3181 acagctatca gttgctcaag gcatatatga tgtcaatatc tccggtctgg taagcacaac
     3241 catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa atcaggaagg
     3301 gatggctgag gtcgcccggt ttattgaaat gaacggctct tttgctgacg agaacagggg
     3361 ctggtgaaat gcagtttaag gtttacacct ataaaagaga gagccgttat cgtctgtttg
     3421 tggatgtaca gagtgatatt attgacacgc ccgggcgacg gatggtgatc cccctggcca
     3481 gtgcacgtct gctgtcagat aaagtctccc gtgaacttta cccggtggtg catatcgggg
     3541 atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggtctcc gttatcgggg
     3601 aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt aacctgatgt
     3661 tctggggaat ataaatgtca ggctccctta tacacagcca gtctgcaggt cgaccatagt
     3721 gactggatat gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt
     3781 aatatattga tatttatatc attttacgtt tctcgttcag ctttcttgta caaagtggtg
     3841 atgggctgca ggaattcgat atcaagctta tcgataccgt cgacctcgag tcatgtaatt
     3901 agttatgtca cgcttacatt cacgccctcc ccccacatcc gctctaaccg aaaaggaagg
     3961 agttagacaa cctgaagtct aggtccctat ttattttttt atagttatgt tagtattaag
     4021 aacgttattt atatttcaaa tttttctttt ttttctgtac agacgcgtgt acgcatgtaa
     4081 cattatactg aaaaccttgc ttgagaaggt tttgggacgc tcgaaggctt taatttgatg
     4141 tcgtaataac cccgccccga aaactgtatt ataagtaaat gcatgtatac taaactcaca
     4201 aattagagct tcaatttaat tatatcagtt attacccggg aatctcggtc gtaatgattt
     4261 ctataatgac gaaaaaaaaa aaattggaaa gaaaaagctt catggccttt ataaaaagga
     4321 actatccaat acctcgccag aaccaagtaa cagtatttta cggggcacaa atcaagaaca
     4381 ataagacagg actgtaaaga tggacgcatt gaactccaaa gaacaacaag agttccaaaa
     4441 agtagtggaa caaaagcaaa tgaaggattt catgcgtttg tactctaatc tggtagaaag
     4501 atgtttcaca gactgtgtca atgacttcac aacatcaaag ctaaccaata aggaacaaac
     4561 atgcatcatg aagtgctcag aaaagttctt gaagcatagc gaacgtgtag ggcagcgttt
     4621 ccaagaacaa aacgctgcct tgggacaagg cttgggccgg tttaaaccat ggtcatagct
     4681 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4741 aaagtgtaaa gcctggggtg cctaatgagt gaggtaactc acattaattg cgttgcgctc
     4801 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4861 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4921 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4981 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5041 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5101 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5161 ccaggcgttc ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5221 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg
     5281 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5341 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5401 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5461 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     5521 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5581 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5641 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5701 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5761 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5821 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5881 tcgttcatcc atagttgcct gactgcccgt cgtgtagata actacgatac gggagggctt
     5941 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6001 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6061 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6121 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6181 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6241 gtgaaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6301 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6361 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6421 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6481 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6541 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6601 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6661 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6721 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6781 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     6841 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtc
//
