LOCUS       Exported                5757 bp ds-DNA     circular SYN 18-JAN-2025
DEFINITION  Plasmid for tagging Gnt1 with moxGFP2 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5757)
  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 GNT1.
FEATURES             Location/Qualifiers
     source          1..5757
                     /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             238..850
                     /codon_start=1
                     /label=GNT1
                     /translation="DIISSTSSLSPLFDFQSSKKVGKLKFASNLMVINPSKEAFDEIVN
                     VMLPKILNKKEKYDMDLINEEMYNLKKIIYKQFIFFRKVRKLFKPEVLVLPFARYGLLT
                     GSLRNPRHYSIIYNDVLGYKTLDNDGNDIPVGLNDSVAYSKYIHFSDYPLAKPWNYPSM
                     KEFECIVKEEDAEDSKLEHQACDLWNSVYASYIQSREICLV"
     CDS             860..1549
                     /codon_start=1
                     /product="oxidation-resistant GFP derivative (Valbuena et 
                     al., 2024)"
                     /label=moxGFP2
                     /note="yeast codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FISTTGKLPVPWPTLVTTLTYGVQVFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGIT"
     3'UTR           1553..2217
                     /label=3' GNT1
     primer_bind     complement(2250..2266)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2734..3537)
                     /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(3538..3753)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     promoter        3851..3955
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3956..4816
                     /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      4987..5575
                     /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 ccaagctgga
      241 cattataagt tctacatcat ctttatcgcc gttatttgat ttccaaagta gtaaaaaggt
      301 tggaaaatta aaatttgctt ctaatttaat ggttattaat ccttcaaagg aggcgtttga
      361 tgaaattgtg aacgttatgc tgcccaagat tttaaacaag aaggaaaagt acgatatgga
      421 tttgatcaat gaagaaatgt acaatttaaa gaagattatt tataaacagt ttattttctt
      481 cagaaaagtc cgaaaacttt tcaaaccaga agtattggtt ttgccatttg caaggtacgg
      541 cttactgacg ggttcattga gaaacccacg gcactattct ataatctata atgacgtttt
      601 gggttataag acattggata atgatggcaa cgatattccg gtaggattaa acgacagtgt
      661 cgcatattct aagtacatac atttttcgga ttatccattg gccaagccct ggaattatcc
      721 atcaatgaag gagtttgaat gtatcgttaa agaagaagac gcggaggatt caaagctaga
      781 gcaccaagct tgtgaccttt ggaactcggt gtacgcttct tacatacagt ccagggaaat
      841 ctgcctcgtt ggaagttcga tggatagcac tgagagcttg ttcacaggtg ttgttcctat
      901 cttggtcgaa ttagacggtg acgtaaacgg tcataagttc agtgttagag gtgaaggtga
      961 aggtgacgct actaacggta aattgacatt gaagtttatt agtactacag gtaaattgcc
     1021 agttccttgg ccaactttgg ttactacatt gacatacggt gttcaagttt tttcaagata
     1081 cccagatcat atgaagcaac atgatttctt taagtctgct atgccagaag gttacgttca
     1141 agaaagaact atcactttta aagatgatgg tacttacaag acaagagcag aggttaagtt
     1201 cgaaggtgac actttggtta acagaatcga attgaagggt atcgatttta aagaagatgg
     1261 taatattttg ggtcataaat tggaatacaa ctacaactca cataacgttt acattacagc
     1321 tgataaacaa aagaatggta ttaaagcaaa cttcaagatc agacataatg ttgaagatgg
     1381 ttctgttcaa ttagcagatc attaccaaca aaacactcca attggtgacg gtccagtttt
     1441 gttaccagat aaccattact tgtctacaca atcaaaattg tctaaagatc caaacgaaaa
     1501 gagagatcat atggttttgt tagaatttgt tactgctgct ggtatcacat gattatgagg
     1561 agaagtgaaa attttcaaaa agtgtaaaaa ttagtatagt cggaaacgca tatatatata
     1621 tcatagtatt gataatgttt ctgccaagat tgtgtccgtt atcgcatttt ctaacgggta
     1681 aggtatgtac taggtgagag cgggatctta tttcaacatg ttgtgcttta atgcttcttt
     1741 attcaacagc ctgtcaacgt catcaccgta ttttgcgtta ccttttccat tttctttgaa
     1801 ggcataatca acagtaattc tgttgttggc gactaactga ttgttcaaag attttattgc
     1861 caaatctgct ttttcaaagt cctcaaaata aacataagca cactttaatt ttccgtttga
     1921 taggtagaaa atttcgggtt cccttatgag cttaccaaat ttattgaaaa ttttcactaa
     1981 ttggtcgcta tctatcgaat ctgcaaggtt ttttatgaac aacttagcaa taggtagaat
     2041 catgtcctta gaaatatttg aaggtaaatt agttgttcct gtagaattgg ttacttgacg
     2101 aacctttatc agtcggtcgt ataatctaac ggtattattc attattttta ttgcatattg
     2161 agcatctcct tgattataaa attcgatgaa tgcgtagcct tggtaagctt gtagaaccta
     2221 gaggatcccc gggtaccgag ctcgaattca ctggccgtcg ttttacaacg tcgtgactgg
     2281 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2341 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2401 gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2461 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     2521 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     2581 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     2641 gcgagacgaa agggcgggta ataactgata taattaaatt gaagctctaa tttgtgagtt
     2701 tagtatacat gcatttactt ataatacagt tttttagttt tgctggccgc atcttctcaa
     2761 atatgcttcc cagcctgctt ttctgtaacg ttcaccctct accttagcat cccttccctt
     2821 tgcaaatagt cctcttccaa caataataat gtcagatcct gtagagacca catcatccac
     2881 ggttctatac tgttgaccca atgcgtctcc cttgtcatct aaacccacac cgggtgtcat
     2941 aatcaaccaa tcgtaacctt catctcttcc acccatgtct ctttgagcaa taaagccgat
     3001 aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga
     3061 tagggagccc ttgcatgaca attctgctaa catcaaaagg cctctaggtt cctttgttac
     3121 ttcttctgcc gcctgcttca aaccgctaac aatacctggg cccaccacac cgtgtgcatt
     3181 cgtaatgtct gcccattctg ctattctgta tacacccgca gagtactgca atttgactgt
     3241 attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa aaattgtact tggcggataa
     3301 tgcctttagc ggcttaactg tgccctccat ggaaaaatca gtcaagatat ccacatgtgt
     3361 ttttagtaaa caaattttgg gacctaatgc ttcaactaac tccagtaatt ccttggtggt
     3421 acgaacatcc aatgaagcac acaagtttgt ttgcttttcg tgcatgatat taaatagctt
     3481 ggcagcaaca ggactaggat gagtagcagc acgttcctta tatgtagctt tcgacatgat
     3541 ttatcttcgt ttcctgcatg tttttgttct gtgcagttgg gttaagaata ctgggcaatt
     3601 tcatgtttct tcaacactac atatgcgtat atataccaat ctaagtctgt gctccttcct
     3661 tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa tttcaaagaa accgaaatca
     3721 aaaaaaagaa taaaaaaaaa atgatgaatt gaattgaaaa gctggcctcg tgatacgcct
     3781 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     3841 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3901 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     3961 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     4021 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     4081 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     4141 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4201 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4261 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4321 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4381 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4441 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4501 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4561 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4621 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4681 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4741 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4801 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     4861 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     4921 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     4981 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5041 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5101 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     5161 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5221 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5281 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5341 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5401 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5461 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5521 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5581 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5641 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5701 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaaga
//
