LOCUS       sequence-71982-e        7634 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FX cloning Saccharomyces cerevisiae expression vector with GAL1 
            promoter and C-terminal 3C protease cleavage site and 10x His tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7634)
  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 7634)
  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: pYEXC3H
FEATURES             Location/Qualifiers
     source          1..7634
                     /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             complement(570..875)
                     /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(672..691)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1217..1876)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1791..1810
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(1930..1936,1937..1954,1955..1960))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(1985..2109)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(2128..2148)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2152..2172)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2199..2222
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2223..2252
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2277..2318
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2328..2345
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2379..2626
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2395..2413)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2703..2720)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2874..3462)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2954..2973)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3633..4424,4425..4493))
                     /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(4589..5392)
                     /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(5393..5613)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      6212..7092
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(7103..7616)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     7510..7529
                     /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 ccttaaccat gagttgaaga gcgacctgca
      541 gactggctgt gtataaggga gcctgacatt tatattcccc agaacatcag gttaatggcg
      601 tttttgatgt cattttcgcg gtggctgaga tcagccactt cttccccgat aacggagacc
      661 ggcacactgg ccatatcggt ggtcatcatg cgccagcttt catccccgat atgcaccacc
      721 gggtaaagtt cacgggagac tttatctgac agcagacgtg cactggccag ggggatcacc
      781 atccgtcgcc cgggcgtgtc aataatatca ctctgtacat ccacaaacag acgataacgg
      841 ctctctcttt tataggtgta aaccttaaac tgcatttcac cagcccctgt tctcgtcagc
      901 aaaagagccg ttcatttcaa taaaccgggc gacctcagcc atcccttcct gattttccgc
      961 tttccagcgt tcggcacgca gacgacgggc ttcattctgc atggttgtgc ttaccagacc
     1021 ggagatattg acatcatata tgccttgagc aactgatagc tgtcgctgtc aactgtcact
     1081 gtaatacgct gcttcatagc atacctcttt ttgacatact tcgggtatac atatcagtat
     1141 atattcttat accgcaaaaa tcagcgcgca aatacgcata ctgttatctg gcttttagta
     1201 agccggatcc acgcgtttac gccccgccct gccactcatc gcagtactgt tgtaattcat
     1261 taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga atcgccagcg
     1321 gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg ggggcgaaga
     1381 agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag ggattggctg
     1441 agacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt tcaccgtaac
     1501 acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg tattcactcc
     1561 agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg tgaacactat
     1621 cccatatcac cagctcaccg tctttcattg ccatacggaa ttccggatga gcattcatca
     1681 ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc tttacggtct
     1741 ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattga gcaactgact
     1801 gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg gtatatccag
     1861 tgattttttt ctccatttta gcttccttag ctcctgaaaa tctcgacgga tcctaactca
     1921 aaatccacac attatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgcgg
     1981 ccgccatagt gactggatat gttgtgtttt acagtattat gtagtctgtt ttttatgcaa
     2041 aatctaattt aatatattga tatttatatc attttacgtt tctcgttcag cttttttgta
     2101 caaacttgtg atatcgaatt cgctagctac ctttctcttc ttttttggat ctacctttct
     2161 cttctttttt ggggatccac tagttcagct cttctgcatt agaagttttg tttcaaggtc
     2221 cacaccatca tcatcaccat catcatcatc attaataatc tagagggccc ttcgaaggta
     2281 agcctatccc taaccctctc ctcggtctcg attctacgcg taccggtcat catcaccatc
     2341 accattgagt ttaaacccgc tgatcctaga gggccgcatc atgtaattag ttatgtcacg
     2401 cttacattca cgccctcccc ccacatccgc tctaaccgaa aaggaaggag ttagacaacc
     2461 tgaagtctag gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat
     2521 atttcaaatt tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa
     2581 aaccttgctt gagaaggttt tgggacgctc gaaggcttta atttgcaagc tgcggccctg
     2641 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctattccgct
     2701 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     2761 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     2821 gcaaaaggcc agcaaaagcc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     2881 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     2941 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3001 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3061 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3121 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3181 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3241 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3301 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3361 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3421 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     3481 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     3541 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3601 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     3661 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     3721 actacgatac gggagcgctt accatctggc cccagtgctg caatgatacc gcgagaccca
     3781 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     3841 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     3901 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg gcattgctac aggcatcgtg
     3961 gtgtcactct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4021 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     4081 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     4141 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     4201 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     4261 agtgtatcac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     4321 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     4381 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     4441 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     4501 ctttttcaat gggtaataac tgatataatt aaattgaagc tctaatttgt gagtttagta
     4561 tacatgcatt tacttataat acagtttttt agttttgctg gccgcatctt ctcaaatatg
     4621 cttcccagcc tgcttttctg taacgttcac cctctacctt agcatccctt ccctttgcaa
     4681 atagtcctct tccaacaata ataatgtcag atcctgtaga gaccacatca tccacggttc
     4741 tatactgttg acccaatgcg tctcccttgt catctaaacc cacaccgggt gtcataatca
     4801 accaatcgta accttcatct cttccaccca tgtctctttg agcaataaag ccgataacaa
     4861 aatctttgtc gcttttcgca atgtcaacag tacccttagt atattctcca gtagataggg
     4921 agcccttgca tgacaattct gctaacatca aaaggcctct aggttccttt gttacttctt
     4981 ctgccgcctg cttcaaaccg ctaacaatac ctgggcccac cacaccgtgt gcattcgtaa
     5041 tgtctgccca ttctgctatt ctgtatacac ccgcagagta ctgcaatttg actgtattac
     5101 caatgtcagc aaattttctg tcttcgaaga gtaaaaaatt gtacttggcg gataatgcct
     5161 ttagcggctt aactgtgccc tccatggaaa aatcagtcaa gatatccaca tgtgttttta
     5221 gtaaacaaat tttgggacct aatgcttcaa ctaactccag taattccttg gtggtacgaa
     5281 catccaatga agcacacaag tttgtttgct tttcgtgcat gatattaaat agcttggcag
     5341 caacaggact aggatgagta gcagcacgtt ccttatatgt agctttcgac atgatttatc
     5401 ttcgtttcct gcaggttttt gttctgtgca gttgggttaa gaatactggg caatttcatg
     5461 tttcttcaac actacatatg cgtatatata ccaatctaag tctgtgctcc ttccttcgtt
     5521 cttccttctg ttcggagatt accgaatcaa aaaaatttca aagaaaccga aatcaaaaaa
     5581 aagaataaaa aaaaaatgat gaattgaatt gaaaagctag cttatcgatg ataagctgtc
     5641 aaagatgaga attaattcca cggactatag actatactag atactccgtc tactgtacga
     5701 tacacttccg ctcaggtcct tgtcctttaa cgaggcctta ccactctttt gttactctat
     5761 tgatccagct cagcaaaggc agtgtgatct aagattctat cttcgcgatg tagtaaaact
     5821 agctagaccg agaaagagac tagaaatgca aaaggcactt ctacaatggc tgccatcatt
     5881 attatccgat gtgacgctgc agcttctcaa tgatattcga atacgctttg aggagataca
     5941 gcctaatatc cgacaaactg ttttacagat ttacgatcgt acttgttacc catcattgaa
     6001 ttttgaacat ccgaacctgg gagttttccc tgaaacagat agtatatttg aacctgtata
     6061 ataatatata gtctagcgct ttacggaaga caatgtatgt atttcggttc ctggagaaac
     6121 tattgcatct attgcatagg taatcttgca cgtcgcatcc ccggttcatt ttctgcgttt
     6181 ccatcttgca cttcaatagc atatctttgt taacgaagca tctgtgcttc attttgtaga
     6241 acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag aatctgagct gcatttttac
     6301 agaacagaaa tgcaacgcga aagcgctatt ttaccaacga agaatctgtg cttcattttt
     6361 gtaaaacaaa aatgcaacgc gacgagagcg ctaatttttc aaacaaagaa tctgagctgc
     6421 atttttacag aacagaaatg caacgcgaga gcgctatttt accaacaaag aatctatact
     6481 tcttttttgt tctacaaaaa tgcatcccga gagcgctatt tttctaacaa agcatcttag
     6541 attacttttt ttctcctttg tgcgctctat aatgcagtct cttgataact ttttgcactg
     6601 taggtccgtt aaggttagaa gaaggctact ttggtgtcta ttttctcttc cataaaaaaa
     6661 gcctgactcc acttcccgcg tttactgatt actagcgaag ctgcgggtgc attttttcaa
     6721 gataaaggca tccccgatta tattctatac cgatgtggat tgcgcatact ttgtgaacag
     6781 aaagtgatag cgttgatgat tcttcattgg tcagaaaatt atgaacggtt tcttctattt
     6841 tgtctctata tactacgtat aggaaatgtt tacattttcg tattgttttc gattcactct
     6901 atgaatagtt cttactacaa tttttttgtc taaagagtaa tactagagat aaacataaaa
     6961 aatgtagagg tcgagtttag atgcaagttc aaggagcgaa aggtggatgg gtaggttata
     7021 tagggatata gcacagagat atatagcaaa gagatacttt tgagcaatgt ttgtggaagc
     7081 ggtattcgca atgggaagct ccaccccggt tgataatcag aaaagcccca aaaacaggaa
     7141 gattgtataa gcaaatattt aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa
     7201 tttttgttaa atcagctcat tttttaacga atagcccgaa atcggcaaaa tcccttataa
     7261 atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttccaaca agagtccact
     7321 attaaagaac gtggactcca acgtcaaagg gcgaaaaagg gtctatcagg gcgatggccc
     7381 actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa
     7441 tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc
     7501 gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt
     7561 cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtgggg
     7621 atcgatccac tagt
//
