LOCUS       Exported                8049 bp ds-DNA     circular SYN 07-JAN-2025
DEFINITION  Plasmid for tagging Pmr1 with msGFP2 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8049)
  AUTHORS   Adam Krahn
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 2, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize with AvrII for integration at PMR1.
FEATURES             Location/Qualifiers
     source          1..8049
                     /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"
     5'UTR           238..678
                     /label=5' PMR1
     CDS             679..1389
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /note="yeast codon-optimized"
                     /translation="MSDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITGGSGGS"
     CDS             1408..4257
                     /codon_start=1
                     /label=PMR1
                     /translation="SDNPFNASLLDEDSNREREILDATAEALSKPSPSLEYCTLSVDEA
                     LEKLDTDKNGGLRSSNEANNRRSLYGPNEITVEDDESLFKKFLSNFIEDRMILLLIGSA
                     VVSLFMGNIDDAVSITLAIFIVVTVGFVQEYRSEKSLEALNKLVPAECHLMRCGQESHV
                     LASTLVPGDLVHFRIGDRIPADIRIIEAIDLSIDESNLTGENEPVHKTSQTIEKSSFND
                     QPNSIVPISERSCIAYMGTLVKEGHGKGIVVGTGTNTSFGAVFEMMNNIEKPKTPLQLT
                     MDKLGKDLSLVSFIVIGMICLVGIIQGRSWLEMFQISVSLAVAAIPEGLPIIVTVTLAL
                     GVLRMAKRKAIVRRLPSVETLGSVNVICSDKTGTLTSNHMTVSKLWCLDSMSNKLNVLS
                     LDKNKKTKNSNGNLKNYLTEDVRETLTIGNLCNNASFSQEHAIFLGNPTDVALLEQLAN
                     FEMPDIRNTVQKVQELPFNSKRKLMATKILNPVDNKCTVYVKGAFERILEYSTSYLKSK
                     GKKTEKLTEAQKATINECANSMASEGLRVFGFAKLTLSDSSTPLTEDLIKDLTFTGLIG
                     MNDPPRPNVKFAIEQLLQGGVHIIMITGDSENTAVNIAKQIGIPVIDPKLSVLSGDKLD
                     EMSDDQLANVIDHVNIFARATPEHKLNIVRALRKRGDVVAMTGDGVNDAPALKLSDIGV
                     SMGRIGTDVAKEASDMVLTDDDFSTILTAIEEGKGIFNNIQNFLTFQLSTSVAALSLVA
                     LSTAFKLPNPLNAMQILWINILMDGPPAQSLGVEPVDHEVMKKPPRKRTDKILTHDVMK
                     RLLTTAACIIVGTVYIFVKEMAEDGKVTARDTTMTFTCFVFFDMFNALACRHNTKSIFE
                     IGFFTNKMFNYAVGLSLLGQMCAIYIPFFQSIFKTEKLGISDILLLLLISSSVFIVDEL
                     RKLWTRKKNEEDSTYFSNV"
     3'UTR           4258..4509
                     /label=3' PMR1
     primer_bind     complement(4542..4558)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(5026..5829)
                     /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(5830..6045)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     promoter        6143..6247
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6248..7108
                     /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      7279..7867
                     /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 ccaagctaga
      241 gttgcgttcc tagcaaggat cggttggtat caattgcacg ttcagctcat ccctagataa
      301 ccacttaaca gcaaagccga tcgttaaact tttgtattct acattctagt tgcatttttt
      361 ttttttttca ttttgttaca tcaagacaag attctctatt taaagaagta ctttttttgt
      421 taacagtgtg gaccctacct atcgttatat cgagatcttg ttccctaggc catcgtacac
      481 tatagccctt ccttcaagca cttaatagaa aaacctcttt ttctacacca tcataatagt
      541 gtttgctcgc cccgttcttt ccattcccat ttataacaat actagtaact ataataatat
      601 ccttacgact gggcaagaca agacgaagca aggccagcac agacgtaagc ttaagtgtaa
      661 gtaaaagata agataattat gagtgatagc actgagagct tgttcacagg tgttgttcct
      721 atcttggtcg aattagacgg tgacgtaaac ggtcataagt tcagtgttag aggtgaaggt
      781 gaaggtgacg ctactaacgg taaattgaca ttgaagttta tttgtactac aggtaaattg
      841 ccagttcctt ggccaacttt ggttactaca ttgacatacg gtgttcaatg tttttcaaga
      901 tacccagatc atatgaagca acatgatttc tttaagtctg ctatgccaga aggttacgtt
      961 caagaaagaa ctatcacttt taaagatgat ggtacttaca agacaagagc agaggttaag
     1021 ttcgaaggtg acactttggt taacagaatc gaattgaagg gtatcgattt taaagaagat
     1081 ggtaatattt tgggtcataa attggaatac aactacaact cacataacgt ttacattaca
     1141 gctgataaac aaaagaatgg tattaaagca aacttcaaga tcagacataa tgttgaagat
     1201 ggttctgttc aattagcaga tcattaccaa caaaacactc caattggtga cggtccagtt
     1261 ttgttaccag ataaccatta cttgtctaca caatcaaaat tgtctaaaga tccaaacgaa
     1321 aagagagatc atatggtttt gttagaattt gttactgctg ctggtatcac aggtggcagt
     1381 ggcggttctg gaggttctgg aggttccagt gacaatccat ttaatgcgag tcttcttgac
     1441 gaggactcaa accgtgagag agaaatacta gatgccacag cagaggccct ttcgaaacca
     1501 agcccttctt tagagtattg tactttatcc gtggacgaag ctctagaaaa actggacact
     1561 gacaaaaacg gtggtttacg atcatctaac gaggccaaca ataggagatc actttatggc
     1621 cccaatgaaa taaccgtaga agatgatgaa agtcttttca agaagttctt gtcaaatttc
     1681 attgaggatc gaatgattct acttttaata ggatccgcag tggtctctct ttttatgggt
     1741 aacattgatg atgctgttag tatcacactg gccattttca tagttgtcac tgtcggtttt
     1801 gtccaagaat ataggtctga aaaatctcta gaagcgttga ataaattggt tcctgctgaa
     1861 tgtcacttaa tgagatgtgg tcaagagagt catgtactgg cttccacctt ggttcctggt
     1921 gatttagtgc acttcagaat aggtgacaga atccccgcag acattagaat tattgaagca
     1981 atcgatttat ccatcgatga aagtaattta actggtgaaa atgaaccggt acataaaacc
     2041 tcacaaacga tcgaaaaatc ttcctttaac gatcagccta attcaattgt accgatttct
     2101 gagagatctt gtatagctta tatgggtaca ttagtcaagg aaggtcatgg taagggtatc
     2161 gtcgtaggaa caggtacaaa cacatccttt ggtgccgttt ttgaaatgat gaataatatt
     2221 gaaaaaccga agactccatt gcagttaaca atggacaaat tgggaaagga cttgtcactg
     2281 gttagcttca tagttattgg tatgatttgt ttagttggta tcatacaagg tagatcttgg
     2341 ttagaaatgt tccaaatatc ggtatcctta gcggttgctg ctattccaga agggttacca
     2401 attattgtca ctgttacttt ggcattgggt gttctgagaa tggccaagcg taaagccatc
     2461 gtgagaaggt taccaagtgt cgaaacttta ggctctgtca acgttatctg ctccgacaaa
     2521 acaggtacac taacctcaaa ccacatgacc gtatctaaac tttggtgctt ggacagtatg
     2581 tccaataagc taaacgtcct ctcattagac aaaaataaga agactaaaaa ttctaatgga
     2641 aatttgaaaa actatttgac tgaagacgtt agggaaactc taactatcgg taatctctgt
     2701 aataatgcat ctttctctca agaacatgcc atatttctgg gaaatcctac tgatgtagct
     2761 cttttagagc aattggcaaa ctttgaaatg cctgatatca gaaacaccgt tcaaaaagtt
     2821 caggaacttc catttaactc gaaaagaaaa ttaatggcaa ccaagattct caaccctgtc
     2881 gacaataagt gtacagttta tgttaaaggt gcatttgaaa gaattcttga gtactccaca
     2941 agttatttga aatcaaaggg taaaaaaact gaaaagttga ctgaagccca aaaagctacg
     3001 ataaatgagt gcgcaaattc tatggcatct gaaggtttgc gtgtctttgg atttgctaaa
     3061 ctaactttgt ctgattcatc aactcctcta accgaagacc taatcaaaga tttaaccttt
     3121 actggtttaa tcggtatgaa tgacccacca agaccgaacg ttaaatttgc catcgaacaa
     3181 ttactacaag gtggtgtcca tattattatg atcactggtg attctgagaa taccgcagta
     3241 aacattgcaa aacaaattgg tattccagtt attgatccaa agctttccgt tttatccggt
     3301 gataaattag atgaaatgtc agatgatcaa ctggccaatg tcatcgacca cgttaatatt
     3361 tttgctcgtg ctacgcctga gcataagtta aacattgttc gtgcattaag aaagaggggt
     3421 gatgtggtag caatgactgg tgatggtgtt aacgacgctc ctgcgttgaa actttcagat
     3481 attggtgttt ctatgggtag aattggtaca gatgtagcca aagaagcctc agatatggtc
     3541 ttaactgatg atgacttcag tactatttta actgccattg aagagggtaa aggtatcttt
     3601 aataatattc agaatttcct gacttttcaa ttgtctactt ctgttgccgc actatcatta
     3661 gttgcactat ctacagcgtt taaactaccc aatccactga acgcaatgca aattctttgg
     3721 ataaatattt taatggatgg gccaccagct caatccttag gtgtggaacc tgttgatcat
     3781 gaagttatga aaaaacctcc aagaaaacgt accgataaaa ttttgaccca tgatgtaatg
     3841 aaacgtttac taaccaccgc ggcctgtatc atcgttggga cagtttacat ttttgttaaa
     3901 gagatggccg aagatggtaa agtaactgct agagatacta ctatgacatt tacttgtttt
     3961 gttttttttg atatgtttaa tgctttggcc tgcagacata acacaaagtc aatcttcgaa
     4021 atcggctttt tcacgaacaa aatgttcaac tacgccgttg gactgtctct gttaggtcaa
     4081 atgtgcgcta tatatatacc atttttccaa agtatcttta aaactgagaa acttggtatc
     4141 tctgatatac tattgttatt gctcatcagc agtagcgttt tcatcgttga tgaattgaga
     4201 aaattgtgga cgaggaaaaa gaatgaagaa gactcaacgt atttctcaaa tgtttgatat
     4261 gtcacatttt gtgcttttat cgtttttcct tccttccctt tatctttcat gaggacgccc
     4321 aaccctattg aggtaaatgt actaattaat gggaacatat gtagatgtat atatgtacat
     4381 atatttacaa cggatactaa gataaacatg tatgggcgac ttttcgtata tacaaagcaa
     4441 tataaaattt tattctttcc ttctttttcc ggactagata taaaatctcc gtaatcagtg
     4501 gttatgattc tagaggatcc ccgggtaccg agctcgaatt cactggccgt cgttttacaa
     4561 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     4621 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     4681 agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     4741 tcacaccgca tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     4801 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     4861 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     4921 tcaccgaaac gcgcgagacg aaagggcggg taataactga tataattaaa ttgaagctct
     4981 aatttgtgag tttagtatac atgcatttac ttataataca gttttttagt tttgctggcc
     5041 gcatcttctc aaatatgctt cccagcctgc ttttctgtaa cgttcaccct ctaccttagc
     5101 atcccttccc tttgcaaata gtcctcttcc aacaataata atgtcagatc ctgtagagac
     5161 cacatcatcc acggttctat actgttgacc caatgcgtct cccttgtcat ctaaacccac
     5221 accgggtgtc ataatcaacc aatcgtaacc ttcatctctt ccacccatgt ctctttgagc
     5281 aataaagccg ataacaaaat ctttgtcgct cttcgcaatg tcaacagtac ccttagtata
     5341 ttctccagta gatagggagc ccttgcatga caattctgct aacatcaaaa ggcctctagg
     5401 ttcctttgtt acttcttctg ccgcctgctt caaaccgcta acaatacctg ggcccaccac
     5461 accgtgtgca ttcgtaatgt ctgcccattc tgctattctg tatacacccg cagagtactg
     5521 caatttgact gtattaccaa tgtcagcaaa ttttctgtct tcgaagagta aaaaattgta
     5581 cttggcggat aatgccttta gcggcttaac tgtgccctcc atggaaaaat cagtcaagat
     5641 atccacatgt gtttttagta aacaaatttt gggacctaat gcttcaacta actccagtaa
     5701 ttccttggtg gtacgaacat ccaatgaagc acacaagttt gtttgctttt cgtgcatgat
     5761 attaaatagc ttggcagcaa caggactagg atgagtagca gcacgttcct tatatgtagc
     5821 tttcgacatg atttatcttc gtttcctgca tgtttttgtt ctgtgcagtt gggttaagaa
     5881 tactgggcaa tttcatgttt cttcaacact acatatgcgt atatatacca atctaagtct
     5941 gtgctccttc cttcgttctt ccttctgttc ggagattacc gaatcaaaaa aatttcaaag
     6001 aaaccgaaat caaaaaaaag aataaaaaaa aaatgatgaa ttgaattgaa aagctggcct
     6061 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     6121 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     6181 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     6241 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     6301 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     6361 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     6421 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     6481 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     6541 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     6601 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     6661 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     6721 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     6781 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     6841 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     6901 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     6961 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     7021 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     7081 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     7141 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     7201 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     7261 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     7321 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     7381 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     7441 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     7501 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     7561 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     7621 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     7681 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     7741 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     7801 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     7861 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     7921 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     7981 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     8041 agcggaaga
//
