LOCUS       sequence-71980-e        7638 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FX cloning Saccharomyces cerevisiae expression vector with GAL1 
            promoter and N-terminal 10x His tag and 3C protease cleavage site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7638)
  AUTHORS   Geertsma ER, Dutzler R
  TITLE     A versatile and efficient high-throughput cloning tool for 
            structural biology.
  JOURNAL   Biochemistry. 2011 Apr 19;50(15):3272-8. Epub 2011 Mar 25.
  PUBMED    21410291
REFERENCE   2  (bases 1 to 7638)
  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: 
            "Biochemistry."; date: "2011-04-19"; volume: "50(15)"; pages: 
            "3272-8. Epub 2011 Mar 25"
COMMENT     Sequence Label: pYEXNH3
FEATURES             Location/Qualifiers
     source          1..7638
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..443
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    2..119
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     414..437
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     475..494
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        475..493
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             523..552
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             553..576
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(625..930)
                     /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     complement(727..746)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1272..1931)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1846..1865
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(1985..1991,1992..2009,2010..2015))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(2040..2164)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2183..2203)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2207..2227)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2281..2322
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2332..2349
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2383..2630
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2399..2417)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2707..2724)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2878..3466)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2958..2977)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3637..4428,4429..4497))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(4593..5396)
                     /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
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(5397..5617)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      6216..7096
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(7107..7620)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     7514..7533
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 acggattaga agccgccgag cgggtgacag ccctccgaag gaagactctc ctccgtgcgt
       61 cctcgtcttc accggtcgcg ttcctgaaac gcagatgtgc ctcgcgccgc actgctccga
      121 acaataaaga ttctacaata ctagctttta tggttatgaa gaggaaaaat tggcagtaac
      181 ctggccccac aaaccttcaa atgaacgaat caaattaaca accataggat gataatgcga
      241 ttagtttttt agccttattt ctggggtaat taatcagcga agcgatgatt tttgatctat
      301 taacagatat ataaatgcaa aaactgcata accactttaa ctaatacttt caacattttc
      361 ggtttgtatt acttcttatt caaatgtaat aaaagtatca acaaaaaatt gttaatatac
      421 ctctatactt taacgtcaag gagaaaaaac cccggatcgg actactagca gctgtaatac
      481 gactcactat agggaatatt aagcttggta ccattaacca tgcaccatca tcatcaccat
      541 catcatcatc atttagaagt tttgtttcaa ggtccaagta gaagagcgag ctgcagactg
      601 gctgtgtata agggagcctg acatttatat tccccagaac atcaggttaa tggcgttttt
      661 gatgtcattt tcgcggtggc tgagatcagc cacttcttcc ccgataacgg agaccggcac
      721 actggccata tcggtggtca tcatgcgcca gctttcatcc ccgatatgca ccaccgggta
      781 aagttcacgg gagactttat ctgacagcag acgtgcactg gccaggggga tcaccatccg
      841 tcgcccgggc gtgtcaataa tatcactctg tacatccaca aacagacgat aacggctctc
      901 tcttttatag gtgtaaacct taaactgcat ttcaccagcc cctgttctcg tcagcaaaag
      961 agccgttcat ttcaataaac cgggcgacct cagccatccc ttcctgattt tccgctttcc
     1021 agcgttcggc acgcagacga cgggcttcat tctgcatggt tgtgcttacc agaccggaga
     1081 tattgacatc atatatgcct tgagcaactg atagctgtcg ctgtcaactg tcactgtaat
     1141 acgctgcttc atagcatacc tctttttgac atacttcggg tatacatatc agtatatatt
     1201 cttataccgc aaaaatcagc gcgcaaatac gcatactgtt atctggcttt tagtaagccg
     1261 gatccacgcg tttacgcccc gccctgccac tcatcgcagt actgttgtaa ttcattaagc
     1321 attctgccga catggaagcc atcacaaacg gcatgatgaa cctgaatcgc cagcggcatc
     1381 agcaccttgt cgccttgcgt ataatatttg cccatggtga aaacgggggc gaagaagttg
     1441 tccatattgg ccacgtttaa atcaaaactg gtgaaactca cccagggatt ggctgagacg
     1501 aaaaacatat tctcaataaa ccctttaggg aaataggcca ggttttcacc gtaacacgcc
     1561 acatcttgcg aatatatgtg tagaaactgc cggaaatcgt cgtggtattc actccagagc
     1621 gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac aagggtgaac actatcccat
     1681 atcaccagct caccgtcttt cattgccata cggaattccg gatgagcatt catcaggcgg
     1741 gcaagaatgt gaataaaggc cggataaaac ttgtgcttat ttttctttac ggtctttaaa
     1801 aaggccgtaa tatccagctg aacggtctgg ttataggtac attgagcaac tgactgaaat
     1861 gcctcaaaat gttctttacg atgccattgg gatatatcaa cggtggtata tccagtgatt
     1921 tttttctcca ttttagcttc cttagctcct gaaaatctcg acggatccta actcaaaatc
     1981 cacacattat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgcggccgcc
     2041 atagtgactg gatatgttgt gttttacagt attatgtagt ctgtttttta tgcaaaatct
     2101 aatttaatat attgatattt atatcatttt acgtttctcg ttcagctttt ttgtacaaac
     2161 ttgtgatatc gaattcgcta gctacctttc tcttcttttt tggatctacc tttctcttct
     2221 tttttgggga tccactagtt cagctcttct gcataataac tatctagagg gcccttcgaa
     2281 ggtaagccta tccctaaccc tctcctcggt ctcgattcta cgcgtaccgg tcatcatcac
     2341 catcaccatt gagtttaaac ccgctgatcc tagagggccg catcatgtaa ttagttatgt
     2401 cacgcttaca ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac
     2461 aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta agaacgttat
     2521 ttatatttca aatttttctt ttttttctgt acagacgcgt gtacgcatgt aacattatac
     2581 tgaaaacctt gcttgagaag gttttgggac gctcgaaggc tttaatttgc aagctgcggc
     2641 cctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctattc
     2701 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     2761 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     2821 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     2881 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     2941 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3001 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3061 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3121 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3181 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3241 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3301 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     3361 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     3421 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     3481 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     3541 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     3601 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     3661 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     3721 gataactacg atacgggagc gcttaccatc tggccccagt gctgcaatga taccgcgaga
     3781 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     3841 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     3901 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttggcattg ctacaggcat
     3961 cgtggtgtca ctctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4021 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4081 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4141 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4201 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     4261 taatagtgta tcacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     4321 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     4381 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     4441 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     4501 cttccttttt caatgggtaa taactgatat aattaaattg aagctctaat ttgtgagttt
     4561 agtatacatg catttactta taatacagtt ttttagtttt gctggccgca tcttctcaaa
     4621 tatgcttccc agcctgcttt tctgtaacgt tcaccctcta ccttagcatc ccttcccttt
     4681 gcaaatagtc ctcttccaac aataataatg tcagatcctg tagagaccac atcatccacg
     4741 gttctatact gttgacccaa tgcgtctccc ttgtcatcta aacccacacc gggtgtcata
     4801 atcaaccaat cgtaaccttc atctcttcca cccatgtctc tttgagcaat aaagccgata
     4861 acaaaatctt tgtcgctttt cgcaatgtca acagtaccct tagtatattc tccagtagat
     4921 agggagccct tgcatgacaa ttctgctaac atcaaaaggc ctctaggttc ctttgttact
     4981 tcttctgccg cctgcttcaa accgctaaca atacctgggc ccaccacacc gtgtgcattc
     5041 gtaatgtctg cccattctgc tattctgtat acacccgcag agtactgcaa tttgactgta
     5101 ttaccaatgt cagcaaattt tctgtcttcg aagagtaaaa aattgtactt ggcggataat
     5161 gcctttagcg gcttaactgt gccctccatg gaaaaatcag tcaagatatc cacatgtgtt
     5221 tttagtaaac aaattttggg acctaatgct tcaactaact ccagtaattc cttggtggta
     5281 cgaacatcca atgaagcaca caagtttgtt tgcttttcgt gcatgatatt aaatagcttg
     5341 gcagcaacag gactaggatg agtagcagca cgttccttat atgtagcttt cgacatgatt
     5401 tatcttcgtt tcctgcaggt ttttgttctg tgcagttggg ttaagaatac tgggcaattt
     5461 catgtttctt caacactaca tatgcgtata tataccaatc taagtctgtg ctccttcctt
     5521 cgttcttcct tctgttcgga gattaccgaa tcaaaaaaat ttcaaagaaa ccgaaatcaa
     5581 aaaaaagaat aaaaaaaaaa tgatgaattg aattgaaaag ctagcttatc gatgataagc
     5641 tgtcaaagat gagaattaat tccacggact atagactata ctagatactc cgtctactgt
     5701 acgatacact tccgctcagg tccttgtcct ttaacgaggc cttaccactc ttttgttact
     5761 ctattgatcc agctcagcaa aggcagtgtg atctaagatt ctatcttcgc gatgtagtaa
     5821 aactagctag accgagaaag agactagaaa tgcaaaaggc acttctacaa tggctgccat
     5881 cattattatc cgatgtgacg ctgcagcttc tcaatgatat tcgaatacgc tttgaggaga
     5941 tacagcctaa tatccgacaa actgttttac agatttacga tcgtacttgt tacccatcat
     6001 tgaattttga acatccgaac ctgggagttt tccctgaaac agatagtata tttgaacctg
     6061 tataataata tatagtctag cgctttacgg aagacaatgt atgtatttcg gttcctggag
     6121 aaactattgc atctattgca taggtaatct tgcacgtcgc atccccggtt cattttctgc
     6181 gtttccatct tgcacttcaa tagcatatct ttgttaacga agcatctgtg cttcattttg
     6241 tagaacaaaa atgcaacgcg agagcgctaa tttttcaaac aaagaatctg agctgcattt
     6301 ttacagaaca gaaatgcaac gcgaaagcgc tattttacca acgaagaatc tgtgcttcat
     6361 ttttgtaaaa caaaaatgca acgcgacgag agcgctaatt tttcaaacaa agaatctgag
     6421 ctgcattttt acagaacaga aatgcaacgc gagagcgcta ttttaccaac aaagaatcta
     6481 tacttctttt ttgttctaca aaaatgcatc ccgagagcgc tatttttcta acaaagcatc
     6541 ttagattact ttttttctcc tttgtgcgct ctataatgca gtctcttgat aactttttgc
     6601 actgtaggtc cgttaaggtt agaagaaggc tactttggtg tctattttct cttccataaa
     6661 aaaagcctga ctccacttcc cgcgtttact gattactagc gaagctgcgg gtgcattttt
     6721 tcaagataaa ggcatccccg attatattct ataccgatgt ggattgcgca tactttgtga
     6781 acagaaagtg atagcgttga tgattcttca ttggtcagaa aattatgaac ggtttcttct
     6841 attttgtctc tatatactac gtataggaaa tgtttacatt ttcgtattgt tttcgattca
     6901 ctctatgaat agttcttact acaatttttt tgtctaaaga gtaatactag agataaacat
     6961 aaaaaatgta gaggtcgagt ttagatgcaa gttcaaggag cgaaaggtgg atgggtaggt
     7021 tatataggga tatagcacag agatatatag caaagagata cttttgagca atgtttgtgg
     7081 aagcggtatt cgcaatggga agctccaccc cggttgataa tcagaaaagc cccaaaaaca
     7141 ggaagattgt ataagcaaat atttaaattg taaacgttaa tattttgtta aaattcgcgt
     7201 taaatttttg ttaaatcagc tcatttttta acgaatagcc cgaaatcggc aaaatccctt
     7261 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttcc aacaagagtc
     7321 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aagggtctat cagggcgatg
     7381 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcag
     7441 taaatcggaa gggtaaacgg atgccccgat ttagagcttg acggggaaag ccggcgaacg
     7501 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     7561 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     7621 ggggatcgat ccactagt
//
