LOCUS       Exported                7265 bp ds-DNA     circular SYN 07-JAN-2025
DEFINITION  Plasmid for tagging Tmn1 with msGFP2 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7265)
  AUTHORS   Adam Krahn
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 2, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize with BsrGI for integration at TMN1.
FEATURES             Location/Qualifiers
     source          1..7265
                     /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           262..1014
                     /label=5`TMN1
     CDS             1015..1083
                     /codon_start=1
                     /label=pre-Ost1
                     /translation="MRQVWFSWIVGLFLCFFNVSSAA"
     CDS             1084..1788
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /note="yeast codon-optimized"
                     /translation="DSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLEF
                     VTAAGITGGSGGS"
     CDS             1807..3743
                     /codon_start=3
                     /label=TMN1
                     /translation="FYLPGVAPTTYRENDNIPLLVNHLTPSMNYQHKDEDGNNVSGDKE
                     NFLYSYDYYYNRFHFCQPEKVEKQPESLGSVIFGDRIYNSPFQLNMLQEKECESLCKTV
                     IPGDDAKFINKLIKNGFFQNWLIDGLPAAREVYDGRTKTSFYGAGFNLGFVQVTQGTDI
                     EATPKGAETTDKDVELETRNDRNMVKTYELPYFANHFDIMIEYHDRGEGNYRVVGVIVE
                     PVSIKRSSPGTCETTGSPLMLDEENDNEVYFTYSVKFNESATSWATRWDKYLHVYDPSI
                     QWFSLINFSLVVVLLSSVVIHSLLRALKSDFARYNELNLDDDFQEDSGWKLNHGDVFRS
                     PSQSLTLSILVGSGVQLFLMVTCSIFFAALGFLSPSSRGSLATVMFILYALFGFVGSYT
                     SMGIYKFFNGPYWKANLILTPLLVPGAILLIIIALNFFLMFVHSSGVIPASTLFFMVFL
                     WFLFSIPLSFAGSLIARKRCHWDEHPTKTNQIARQIPFQPWYLKTIPATLIAGIFPFGS
                     IAVELYFIYTSLWFNKIFYMFGFLFFSFLLLTLTSSLVTILITYHSLCLENWKWQWRGF
                     IIGGAGCALYVFIHSILFTKFKLGGFTTIVLYVGYSSVISLLCCLVTGSIGFISSMLFV
                     RKIYSSIKVD"
     primer_bind     complement(3758..3774)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4242..5045)
                     /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(5046..5261)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     promoter        5359..5463
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5464..6324
                     /codon_start=1
                     /label=Amp-R
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6495..7083
                     /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 atgcctgcag gtcgactcta gatcatccca gcctccatct tcatcggcaa gcagtgaatc
      301 taaatcaaaa tcgaagaaga agaaaatagg tcgtggtttt gtcgttttaa ttggtcttgc
      361 actagcatta gggacagttt ctgtccttgg tattgcaggc gtcattcttg cctacgtgtt
      421 taaggatcca gagggtgatt ataagcccat taaaccgcga attgatgaga atgaaatgct
      481 tgataccgtc ccacccgaaa aacttatgaa gtttgtctaa tcggggttgt attatataat
      541 atataactac ttaaaataat gaacacttaa tataataaac aaaaataaaa taggaaaatc
      601 ctaagttgat caaaacttaa taaaaattac tacttacact ggaatgttag tagtaaattc
      661 aacacgtatt ggaagtcact tggaatgtta ccaatactgc ctgagaatga gtgattttaa
      721 cttataatac aaaagtccgg gttacagccc tttaaatttt tttcacggca atttttcttt
      781 acttggtgaa gccatgaaaa atgtcatcgg ttcgaagaaa taggcctaat gataaacaaa
      841 aaggtaattc aaggtttttg aactgctttg ttttttttat ttgtatcttt ttttggtacg
      901 ttagttaagt taatttttgg ctccttgtcc agcttttcct tcatctgcta ccgttatatt
      961 ttcagatatt aatttagtag cagagagcaa gtaccgaaag gaaaacaaaa cagaatgagg
     1021 caggtttggt tctcttggat tgtgggattg ttcctatgtt ttttcaacgt gtcttctgct
     1081 gccgatagca ctgagagctt gttcacaggt gttgttccta tcttggtcga attagacggt
     1141 gacgtaaacg gtcataagtt cagtgttaga ggtgaaggtg aaggtgacgc tactaacggt
     1201 aaattgacat tgaagtttat ttgtactaca ggtaaattgc cagttccttg gccaactttg
     1261 gttactacat tgacatacgg tgttcaatgt ttttcaagat acccagatca tatgaagcaa
     1321 catgatttct ttaagtctgc tatgccagaa ggttacgttc aagaaagaac tatcactttt
     1381 aaagatgatg gtacttacaa gacaagagca gaggttaagt tcgaaggtga cactttggtt
     1441 aacagaatcg aattgaaggg tatcgatttt aaagaagatg gtaatatttt gggtcataaa
     1501 ttggaataca actacaactc acataacgtt tacattacag ctgataaaca aaagaatggt
     1561 attaaagcaa acttcaagat cagacataat gttgaagatg gttctgttca attagcagat
     1621 cattaccaac aaaacactcc aattggtgac ggtccagttt tgttaccaga taaccattac
     1681 ttgtctacac aatcaaaatt gtctaaagat ccaaacgaaa agagagatca tatggttttg
     1741 ttagaatttg ttactgctgc tggtatcaca ggtggcagtg gcggttctgg aggctcagga
     1801 gggagcttct atttaccagg tgtagcgcct accacttaca gggaaaatga caacatccca
     1861 ttgctcgtta atcatttaac tccatcaatg aattatcaac acaaagatga ggacggtaat
     1921 aacgtttctg gcgataaaga aaatttcttg tactcatacg actactatta caatagattc
     1981 catttttgtc agcctgaaaa agtcgaaaag caacctgaat ccttgggatc cgttatattt
     2041 ggtgatcgaa tttataattc accattccaa ttgaatatgt tacaagaaaa agagtgtgaa
     2101 tccctgtgta agaccgtaat tccgggagat gatgctaaat tcatcaataa attgataaaa
     2161 aatggcttct tccaaaactg gcttattgat ggattaccgg ctgcaagaga ggtttatgat
     2221 ggcagaacta agactagttt ttatggtgct ggcttcaacc ttggttttgt acaggtaact
     2281 caaggaaccg acattgaggc tactccaaaa ggggcagaaa ccaccgataa ggatgtggaa
     2341 ttggaaacac gtaatgatcg taacatggtg aaaacttacg aattgcctta ctttgccaac
     2401 catttcgata ttatgattga atatcacgat cgtggtgaag gtaactatcg cgtagttggt
     2461 gttatcgtcg aaccagtttc cataaaacgt tccagtccag gtacatgtga aacgactggc
     2521 agcccattga tgctagacga agaaaatgat aatgaagttt attttactta ttccgttaaa
     2581 ttcaatgaat cagcgacttc ttgggccacc aggtgggaca aatatttaca cgtatatgat
     2641 ccttctattc aatggttttc tttgataaat ttctccttag ttgttgtctt attgtcatct
     2701 gttgttatcc attcgctatt gcgcgcactg aaaagtgatt tcgcacgtta caacgagtta
     2761 aatttggacg acgattttca agaagattca ggttggaaat tgaatcatgg tgatgttttt
     2821 cgttcgccaa gtcaatcgct aaccctttca attttggttg gttcaggagt tcaattattt
     2881 ttgatggtaa cttgcagtat cttcttcgct gcattgggtt tcctatcacc tagttctaga
     2941 ggttccttag ccactgttat gtttatctta tatgcattat tcgggtttgt cggctcttat
     3001 acatctatgg gcatctacaa atttttcaat gggccatact ggaaggcaaa cttgatattg
     3061 actccacttc tagttcctgg tgctattctg ctaataatta ttgcgttgaa ctttttttta
     3121 atgtttgttc attcttctgg tgtcatacca gcaagtactc tgttttttat ggtttttcta
     3181 tggtttttgt tctcaattcc attatcattt gctggttcat tgattgccag gaaaagatgt
     3241 cactgggatg aacatccaac caagacaaac caaatcgcaa gacaaatacc cttccagcct
     3301 tggtatttaa agactatacc agctactttg attgctggta tctttccatt tggctccatc
     3361 gccgttgagt tgtatttcat atacacaagt ttatggttta ataagatttt ttacatgttt
     3421 ggtttcctct ttttctcttt cctcctatta actttgacaa gttcactcgt cactattttg
     3481 atcacttatc attccttatg cctggagaac tggaagtggc aatggagagg attcataatt
     3541 ggaggtgccg gttgtgcttt gtatgtgttt attcattcta ttttattcac caaatttaag
     3601 ttgggtggat tcacgactat tgtcttatac gtgggatact cctcagtcat ttcattactc
     3661 tgctgtttgg taacggggtc aattggcttt atcagcagta tgctctttgt tagaaaaatc
     3721 tactcatcca ttaaagttga ctagcgagct cgaattcact ggccgtcgtt ttacaacgtc
     3781 gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg
     3841 ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
     3901 tgaatggcga atggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3961 accgcatatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     4021 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     4081 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     4141 cgaaacgcgc gagacgaaag ggcgggtaat aactgatata attaaattga agctctaatt
     4201 tgtgagttta gtatacatgc atttacttat aatacagttt tttagttttg ctggccgcat
     4261 cttctcaaat atgcttccca gcctgctttt ctgtaacgtt caccctctac cttagcatcc
     4321 cttccctttg caaatagtcc tcttccaaca ataataatgt cagatcctgt agagaccaca
     4381 tcatccacgg ttctatactg ttgacccaat gcgtctccct tgtcatctaa acccacaccg
     4441 ggtgtcataa tcaaccaatc gtaaccttca tctcttccac ccatgtctct ttgagcaata
     4501 aagccgataa caaaatcttt gtcgctcttc gcaatgtcaa cagtaccctt agtatattct
     4561 ccagtagata gggagccctt gcatgacaat tctgctaaca tcaaaaggcc tctaggttcc
     4621 tttgttactt cttctgccgc ctgcttcaaa ccgctaacaa tacctgggcc caccacaccg
     4681 tgtgcattcg taatgtctgc ccattctgct attctgtata cacccgcaga gtactgcaat
     4741 ttgactgtat taccaatgtc agcaaatttt ctgtcttcga agagtaaaaa attgtacttg
     4801 gcggataatg cctttagcgg cttaactgtg ccctccatgg aaaaatcagt caagatatcc
     4861 acatgtgttt ttagtaaaca aattttggga cctaatgctt caactaactc cagtaattcc
     4921 ttggtggtac gaacatccaa tgaagcacac aagtttgttt gcttttcgtg catgatatta
     4981 aatagcttgg cagcaacagg actaggatga gtagcagcac gttccttata tgtagctttc
     5041 gacatgattt atcttcgttt cctgcatgtt tttgttctgt gcagttgggt taagaatact
     5101 gggcaatttc atgtttcttc aacactacat atgcgtatat ataccaatct aagtctgtgc
     5161 tccttccttc gttcttcctt ctgttcggag attaccgaat caaaaaaatt tcaaagaaac
     5221 cgaaatcaaa aaaaagaata aaaaaaaaat gatgaattga attgaaaagc tggcctcgtg
     5281 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     5341 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     5401 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     5461 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     5521 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     5581 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     5641 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     5701 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     5761 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     5821 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     5881 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     5941 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     6001 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     6061 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     6121 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     6181 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     6241 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     6301 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     6361 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     6421 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     6481 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6541 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6601 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     6661 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     6721 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     6781 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6841 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6901 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6961 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     7021 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     7081 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     7141 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     7201 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     7261 gaaga
//
