LOCUS       Exported                5577 bp ds-DNA     circular SYN 18-JAN-2025
DEFINITION  Plasmid for tagging Kex2 with FRBx2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5577)
  AUTHORS   Adam Krahn
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 2, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize with BstXI for integration at KEX2.
FEATURES             Location/Qualifiers
     source          1..5577
                     /organism="synthetic DNA construct"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             244..1013
                     /codon_start=3
                     /label=KEX2
                     /translation="HWGENGVGDWKIKVKTTENGHRIDFHSWRLKLFGESIDSSKTETF
                     VFGNDKEEVEPAATESTVSQYSASSTSISISATSTSSISIGVETSAIPQTTTASTDPDS
                     DPNTPKKLSSPRQAMHYFLTIFLIGATFLVLYFMFFMKSRRRIRRSRAETYEFDIIDTD
                     SEYDSTLDNGTSGITEPEEVEDFDFDLSDEDHLASLSSSENGDAEHTIDSVLTNENPFS
                     DPIKQKFPNDANAESASNKLQELQPDVPPSSGRS"
     CDS             1023..1064
                     /codon_start=1
                     /label=GSS linker
                     /translation="GSSGSSGSSGSSAT"
     CDS             1077..1355
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             1362..1403
                     /codon_start=1
                     /label=GSS linker
                     /translation="GSSGSSGSSGSSAT"
     CDS             1416..1694
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK"
     CDS             1701..1727
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     3'UTR           1740..2053
                     /label=3' KEX2
     primer_bind     complement(2070..2086)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2554..3357)
                     /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(3358..3578)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     promoter        3671..3775
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3776..4636
                     /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"
     rep_origin      4807..5395
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number colE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 atgcacattg gggtgagaac ggcgtaggtg attggaaaat caaggttaag acaacagaaa
      301 atggacacag gattgacttc cacagttgga ggctgaagct ctttggggaa tccattgatt
      361 catctaaaac agaaactttc gtctttggaa acgataaaga ggaggttgaa ccagctgcta
      421 cagaaagtac cgtatcacaa tattctgcca gttcaacttc tatttccatc agcgctactt
      481 ctacatcttc tatctcaatt ggtgtggaaa cgtcggccat tccccaaacg actactgcga
      541 gtaccgatcc tgattctgat ccaaacactc ctaaaaaact ttcctctcct aggcaagcca
      601 tgcattattt tttaacaata tttttgattg gcgccacatt tttggtgtta tacttcatgt
      661 tttttatgaa atcaaggaga aggatcagaa ggtcaagagc ggaaacgtat gaattcgata
      721 tcattgatac agactctgag tacgattcta ctttggacaa tggaacttcc ggaattactg
      781 agcccgaaga ggttgaggac ttcgattttg atttgtccga tgaagaccat cttgcaagtt
      841 tgtcttcatc agaaaacggt gatgctgaac atacaattga tagtgtacta acaaacgaaa
      901 atccatttag tgaccctata aagcaaaagt tcccaaatga cgccaacgca gaatctgctt
      961 ccaataaatt acaagaatta cagcctgatg ttcctccatc ttccggacga tcgacggatc
     1021 caggttcctc tgggtcttcc ggttcctcag ggtcgtcggc caccatggct tctagaatcc
     1081 tctggcatga gatgtggcat gaaggcctgg aagaggcatc tcgtttgtac tttggggaaa
     1141 ggaacgtgaa aggcatgttt gaggtgctgg agcccttgca tgctatgatg gaacggggcc
     1201 cccagactct gaaggaaaca tcctttaatc aggcctatgg tcgagattta atggaggccc
     1261 aagagtggtg caggaagtac atgaaatcag ggaatgtcaa ggacctcacc caagcctggg
     1321 acctctatta tcatgtgttc cgacgaatct caaagactag tggttcctct gggtcttccg
     1381 gttcctcagg gtcgtcggcc accatggctt ctagaatcct ctggcatgag atgtggcatg
     1441 aaggcctgga agaggcatct cgtttgtact ttggggaaag gaacgtgaaa ggcatgtttg
     1501 aggtgctgga gcccttgcat gctatgatgg aacggggccc ccagactctg aaggaaacat
     1561 cctttaatca ggcctatggt cgagatttaa tggaggccca agagtggtgc aggaagtaca
     1621 tgaaatcagg gaatgtcaag gacctcaccc aagcctggga cctctattat catgtgttcc
     1681 gacgaatctc aaagactagt tatccgtacg acgtaccaga ctacgcataa agcggccgct
     1741 tcgatatgta cagaaagctt caaattacaa aatagcattt ttttcttata gattataata
     1801 ctctctcata cgtatacgta tatgtgtata tgatatataa acaaacatta atatcctatt
     1861 ccttccgttt gaaatcccta tgatgtactt tgcattgttt gcacccgcga ataaaatgaa
     1921 aactccgaac cgatatatca agcacataaa aggggagggt ccaattaatg catatttaag
     1981 accacagctg aataacttta aaacggcaga caaaacaaaa aataggtcga ataaacctta
     2041 cctgcctaga aggtaccgag ctcgaattca ctggccgtcg ttttacaacg tcgtgactgg
     2101 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2161 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2221 gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2281 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     2341 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     2401 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     2461 gcgagacgaa agggcgggta ataactgata taattaaatt gaagctctaa tttgtgagtt
     2521 tagtatacat gcatttactt ataatacagt tttttagttt tgctggccgc atcttctcaa
     2581 atatgcttcc cagcctgctt ttctgtaacg ttcaccctct accttagcat cccttccctt
     2641 tgcaaatagt cctcttccaa caataataat gtcagatcct gtagagacca catcatccac
     2701 ggttctatac tgttgaccca atgcgtctcc cttgtcatct aaacccacac cgggtgtcat
     2761 aatcaaccaa tcgtaacctt catctcttcc acccatgtct ctttgagcaa taaagccgat
     2821 aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga
     2881 tagggagccc ttgcatgaca attctgctaa catcaaaagg cctctaggtt cctttgttac
     2941 ttcttctgcc gcctgcttca aaccgctaac aatacctggg cccaccacac cgtgtgcatt
     3001 cgtaatgtct gcccattctg ctattctgta tacacccgca gagtactgca atttgactgt
     3061 attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa aaattgtact tggcggataa
     3121 tgcctttagc ggcttaactg tgccctccat ggaaaaatca gtcaagatat ccacatgtgt
     3181 ttttagtaaa caaattttgg gacctaatgc ttcaactaac tccagtaatt ccttggtggt
     3241 acgaacatcc aatgaagcac acaagtttgt ttgcttttcg tgcatgatat taaatagctt
     3301 ggcagcaaca ggactaggat gagtagcagc acgttcctta tatgtagctt tcgacatgat
     3361 ttatcttcgt ttcctgcatg tttttgttct gtgcagttgg gttaagaata ctgggcaatt
     3421 tcatgtttct tcaacactac atatgcgtat atataccaat ctaagtctgt gctccttcct
     3481 tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa tttcaaagaa accgaaatca
     3541 aaaaaaagaa taaaaaaaaa atgatgaatt gaattgaaaa gctggcctcg tgatacgcct
     3601 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     3661 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3721 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     3781 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     3841 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     3901 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     3961 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4021 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4081 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4141 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4201 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4261 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4321 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4381 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4441 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4501 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4561 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4621 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     4681 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     4741 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     4801 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     4861 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     4921 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     4981 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5041 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5101 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5161 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5221 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5281 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5341 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5401 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5461 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5521 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaaga
//
