LOCUS       sequence-402855-        7896 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast expression vector; To express proteins under control of TDH3 
            promoter and terminator using leucine selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7896)
  AUTHORS   Park EJ, Grabinska KA, Guan Z, Stranecky V, Hartmannova H, Hodanova 
            K, Baresova V, Sovova J, Jozsef L, Ondruskova N, Hansikova H, Honzik
            T, Zeman J, Hulkova H, Wen R, Kmoch S, Sessa WC
  TITLE     Mutation of Nogo-B receptor, a subunit of cis-prenyltransferase, 
            causes a congenital disorder of glycosylation.
  JOURNAL   Cell Metab. 2014 Sep 2;20(3):448-57. doi: 
            10.1016/j.cmet.2014.06.016. Epub 2014 Jul 24.
  PUBMED    25066056
REFERENCE   2  (bases 1 to 7896)
  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"; doi: 
            "10.1016/j.cmet.2014.06.016"; journalName: "Cell Metab"; date: 
            "2014-09-2- 2"; volume: "20"; issue: "3"; pages: "448-57"
COMMENT     Sequence Label: pKG-GW1
FEATURES             Location/Qualifiers
     source          1..7896
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        54..720
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     590..613
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     protein_bind    723..847
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(872..877,878..895,896..902)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             956..1615
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1022..1041)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1957..2262
                     /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     2141..2160
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2303..2427)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2632..2649)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2803..3391)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2883..2902)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3562..4353,4354..4422))
                     /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     4185..4204
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4423..4527)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4595..4613
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4651..4673)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4773..4792
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      4809..5791
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        5792..6197
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             6198..7292
                     /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"
ORIGIN
        1 ggatccggta gaatcatttt gaataaaaaa cacgcttttt cagttcgagt ttatcattat
       61 caatactgcc atttcaaaga atacgtaaat aattaatagt agtgattttc ctaactttat
      121 ttagtcaaaa aattagcctt ttaattctgc tgtaacccgt acatgcccaa aatagggggc
      181 gggttacaca gaatatataa catcgtaggt gtctgggtga acagtttatt cctggcatcc
      241 actaaatata atggagcccg ctttttaagc tggcatccag aaaaaaaaag aatcccagca
      301 ccaaaatatt gttttcttca ccaaccatca gttcataggt ccattctctt agcgcaacta
      361 cagagaacag gggcacaaac aggcaaaaaa cgggcacaac ctcaatggag tgatgcaacc
      421 tgcctggagt aaatgatgac acaaggcaat tgacccacgc atgtatctat ctcattttct
      481 tacaccttct attaccttct gctctctctg atttggaaaa agctgaaaaa aaaggttgaa
      541 accagttccc tgaaattatt cccctacttg actaataagt atataaagac ggtaggtatt
      601 gattgtaatt ctgtaaatct atttcttaaa cttcttaaat tctactttta tagttagtct
      661 tttttttagt tttaaaacac caagaactta gtttcgaata aacacacata aacagaatta
      721 tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa tgatataaat atcaatatat
      781 taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa catatccagt
      841 cactatggcg gccgcattag gcaccccagg ctttacactt tatgcttccg gctcgtataa
      901 tgtgtggatt ttgagttagg atccgtcgag attttcagga gctaaggaag ctaaaatgga
      961 gaaaaaaatc actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt
     1021 tgaggcattt cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac
     1081 ggccttttta aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat
     1141 tcttgcccgc ctgatgaatg ctcatccgga attccgtatg gcaatgaaag acggtgagct
     1201 ggtgatatgg gatagtgttc acccttgtta caccgttttc catgagcaaa ctgaaacgtt
     1261 ttcatcgctc tggagtgaat accacgacga tttccggcag tttctacaca tatattcgca
     1321 agatgtggcg tgttacggtg aaaacctggc ctatttccct aaagggttta ttgagaatat
     1381 gtttttcgtc tcagccaatc cctgggtgag tttcaccagt tttgatttaa acgtggccaa
     1441 tatggacaac ttcttcgccc ccgttttcac catgggcaaa tattatacgc aaggcgacaa
     1501 ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt tgtgatggct tccatgtcgg
     1561 cagaatgctt aatgaattac aacagtactg cgatgagtgg cagggcgggg cgtaaacgcg
     1621 tggatccggc ttactaaaag ccagataaca gtatgcgtat ttgcgcgctg atttttgcgg
     1681 tataagaata tatactgata tgtatacccg aagtatgtca aaaagaggta tgctatgaag
     1741 cagcgtatta cagtgacagt tgacagcgac agctatcagt tgctcaaggc atatatgatg
     1801 tcaatatctc cggtctggta agcacaacca tgcagaatga agcccgtcgt ctgcgtgccg
     1861 aacgctggaa agcggaaaat caggaaggga tggctgaggt cgcccggttt attgaaatga
     1921 acggctcttt tgctgacgag aacaggggct ggtgaaatgc agtttaaggt ttacacctat
     1981 aaaagagaga gccgttatcg tctgtttgtg gatgtacaga gtgatattat tgacacgccc
     2041 gggcgacgga tggtgatccc cctggccagt gcacgtctgc tgtcagataa agtctcccgt
     2101 gaactttacc cggtggtgca tatcggggat gaaagctggc gcatgatgac caccgatatg
     2161 gccagtgtgc cggtctccgt tatcggggaa gaagtggctg atctcagcca ccgcgaaaat
     2221 gacatcaaaa acgccattaa cctgatgttc tggggaatat aaatgtcagg ctcccttata
     2281 cacagccagt ctgcaggtcg accatagtga ctggatatgt tgtgttttac agtattatgt
     2341 agtctgtttt ttatgcaaaa tctaatttaa tatattgata tttatatcat tttacgtttc
     2401 tcgttcagct ttcttgtaca aagtggtgat tcgacttggt tgaacacgtt gccaaggctt
     2461 aagtgaattt actttaaatc ttgcatttaa ataaattttc tttttatagc tttatgactt
     2521 agtttcaatt tatatactat tttaatgaca ttttcgattc ggatccgtgc attaatgaat
     2581 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     2641 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     2701 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     2761 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     2821 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2881 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2941 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     3001 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     3061 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3121 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3181 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3241 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3301 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3361 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     3421 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3481 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3541 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     3601 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     3661 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     3721 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     3781 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     3841 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     3901 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3961 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     4021 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     4081 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     4141 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     4201 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     4261 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     4321 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     4381 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     4441 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     4501 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     4561 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     4621 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     4681 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     4741 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
     4801 gcaccatact agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
     4861 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
     4921 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
     4981 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
     5041 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
     5101 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
     5161 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
     5221 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
     5281 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
     5341 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     5401 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     5461 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     5521 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     5581 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     5641 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     5701 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     5761 gttctacaaa atgaagcaca gatgcttcgt taactgtggg aatactcagg tatcgtaaga
     5821 tgcaagagtt cgaatctctt agcaaccatt atttttttcc tcaacataac gagaacacac
     5881 aggggcgcta tcgcacagaa tcaaattcga tgactggaaa ttttttgtta atttcagagg
     5941 tcgcctgacg catatacctt tttcaactga aaaattggga gaaaaaggaa aggtgagagc
     6001 gccggaaccg gcttttcata tagaatagag aagcgttcat gactaaatgc ttgcatcaca
     6061 atacttgaag ttgacaatat tatttaagga cctattgttt tttccaatag gtggttagca
     6121 atcgtcttac tttctaactt ttcttacctt ttacatttca gcaatatata tatatatttc
     6181 aaggatatac cattctaatg tctgccccta agaagatcgt cgttttgcca ggtgaccacg
     6241 ttggtcaaga aatcacagcc gaagccatta aggttcttaa agctatttct gatgttcgtt
     6301 ccaatgtcaa gttcgatttc gaaaatcatt taattggtgg tgctgctatc gatgctacag
     6361 gtgttccact tccagatgag gcgctggaag cctccaagaa ggctgatgcc gttttgttag
     6421 gtgctgtggg tggtcctaaa tggggtaccg gtagtgttag acctgaacaa ggtttactaa
     6481 aaatccgtaa agaacttcaa ttgtacgcca acttaagacc atgtaacttt gcatccgact
     6541 ctcttttaga cttatctcca atcaagccac aatttgctaa aggtactgac ttcgttgttg
     6601 tcagagaatt agtgggaggt atttactttg gtaagagaaa ggaagacgat ggtgatggtg
     6661 tcgcttggga tagtgaacaa tacaccgttc cagaagtgca aagaatcaca agaatggccg
     6721 ctttcatggc cctacaacat gagccaccat tgcctatttg gtccttggat aaagctaatg
     6781 ttttggcctc ttcaagatta tggagaaaaa ctgtggagga aaccatcaag aacgaattcc
     6841 ctacattgaa ggttcaacat caattgattg attctgccgc catgatccta gttaagaacc
     6901 caacccacct aaatggtatt ataatcacca gcaacatgtt tggtgatatc atctccgatg
     6961 aagcctccgt tatcccaggt tccttgggtt tgttgccatc tgcgtccttg gcctctttgc
     7021 cagacaagaa caccgcattt ggtttgtacg aaccatgcca cggttctgct ccagatttgc
     7081 caaagaataa ggtcaaccct atcgccacta tcttgtctgc tgcaatgatg ttgaaattgt
     7141 cattgaactt gcctgaagaa ggtaaggcca ttgaagatgc agttaaaaag gttttggatg
     7201 caggtatcag aactggtgat ttaggtggtt ccaacagtac caccgaagtc ggtgatgctg
     7261 tcgccgaaga agttaagaaa atccttgctt aaaaagattc tcttttttta tgatatttgt
     7321 acataaactt tataaatgaa attcataata gaaacgacac gaaattacaa aatggaatat
     7381 gttcataggg tagacgaaac tatatacgca atctacatac atttatcaag aaggagaaaa
     7441 aggaggatgt aaaggaatac aggtaagcaa attgatacta atggctcaac gtgataagga
     7501 aaaagaattg cactttaaca ttaatattga caaggaggag ggcaccacac aaaaagttag
     7561 gtgtaacaga aaatcatgaa actatgattc ctaatttata tattggagga ttttctctaa
     7621 aaaaaaaaaa atacaacaaa taaaaaacac tcaatgacct gaccatttga tggagtttaa
     7681 gtcaatacct tcttgaacca tttcccataa tggtgaaagt tccctcaaga attttactct
     7741 gtcagaaacg gccttaacga cgtaccggat ctatgcggtg tgaaataccg cacagatgcg
     7801 taaggagaaa ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag
     7861 ggcgatcggt gcgggcctct tcgctattac gccagc
//
