LOCUS       Exported                7071 bp ds-DNA     circular SYN 06-JAN-2025
DEFINITION  Plasmid for tagging Gga2 with msGFPx3 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7071)
  AUTHORS   Glick Lab
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 2, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize at AflII for integration at GGA2.
FEATURES             Location/Qualifiers
     source          1..7071
                     /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..989
                     /codon_start=3
                     /label=GGA2
                     /translation="PLQQSSGALTNEINLIDFNDLDEAPSQGNNNTNGTGTPAAAETSV
                     NDLLGDLTDLSISNPSTANQASFGLGGDIVLGSSQPAPPVTTTNNSNNTLDLLGLSTPQ
                     SPTNSQAVNSSGFDLLMGFNPTTGTTTAPARTLVNQSPNLKIEFEISRESNSVIRIKSF
                     FTNLSSSPISNLVFLLAVPKSMSLKLQPQSSNFMIGNAKDGISQEGTIENAPANPSKAL
                     KVKWKVNYSVNSTQAEETAVFTLPNV"
     CDS             999..1034
                     /codon_start=1
                     /label=GSS linker
                     /translation="GSSGSSGSSGAS"
     CDS             1035..1748
                     /codon_start=1
                     /product="monomeric superfolder GFP"
                     /label=msGFP
                     /note="mammalian codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGSSGSSG"
     CDS             1755..2468
                     /codon_start=1
                     /product="monomeric superfolder GFP"
                     /label=msGFP
                     /note="mammalian codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGSSGSSG"
     CDS             2475..3191
                     /codon_start=1
                     /product="monomeric superfolder GFP"
                     /label=msGFP
                     /note="mammalian codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGSSGSSG"
     3'UTR           3201..3449
                     /label=3' GGA2
     primer_bind     complement(3564..3580)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4048..4851)
                     /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(4852..5072)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     promoter        5165..5269
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5270..6130
                     /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      6301..6889
                     /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 atgctccgtt acaacaatct tctggtgctt taacgaatga aatcaactta atcgatttca
      301 atgatttgga cgaagcgccc tctcaaggca acaacaatac caacggcacg ggtacaccag
      361 cagcagcgga aacatctgtc aatgacttat tgggcgattt gaccgattta tctatttcga
      421 acccttcaac cgctaaccaa gcatcctttg gtctgggagg cgatatcgtg ttggggtcct
      481 ctcaacccgc accaccagtt actactacta acaactcgaa caacacttta gatcttttgg
      541 gactttcaac tcctcaatcg ccaactaatt cccaagcagt aaacagtagt gggttcgatc
      601 ttttgatggg atttaatcca acaacaggta ctactactgc gcccgcaaga accctagtca
      661 atcagtctcc taacttgaag attgagttcg aaatatctag ggagtcaaac tcagttataa
      721 ggataaaatc gttctttaca aacttaagtt cgtcaccgat ctccaactta gtgttcttat
      781 tagcagtccc taagtcaatg tctttgaaat tgcaacccca atcgagtaac tttatgattg
      841 gcaacgctaa agatggtatc tctcaagaag gtacaattga aaatgcgcca gctaaccctt
      901 caaaggcttt gaaagtcaag tggaaggtca actattctgt caactccacc caagctgaag
      961 aaactgctgt ttttacgtta cctaatgtaa cggatccagg aagttcgggg tccagcggtt
     1021 cttcaggcgc tagcatggat agcactgaga gcctgttcac cggggtggtg cccatcctgg
     1081 tcgagctgga cggcgacgta aacggccaca agttcagcgt gcgcggcgag ggcgagggcg
     1141 atgccaccaa cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1201 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1261 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1321 gcaccatcac cttcaaggac gacggcacct acaagacccg cgccgaggtg aagttcgagg
     1381 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1441 tcctggggca caagctggag tacaacttca acagccacaa cgtctatatc accgccgaca
     1501 agcagaagaa cggcatcaag gccaacttca agatccgcca caacgtggag gacggcagcg
     1561 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1621 ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac gagaagcgcg
     1681 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactcacggc agctccggct
     1741 ccagcgggtc tagcatggat agcactgaga gcctgttcac cggggtggtg cccatcctgg
     1801 tcgagctgga cggcgacgta aacggccaca agttcagcgt gcgcggcgag ggcgagggcg
     1861 atgccaccaa cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1921 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1981 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     2041 gcaccatcac cttcaaggac gacggcacct acaagacccg cgccgaggtg aagttcgagg
     2101 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     2161 tcctggggca caagctggag tacaacttca acagccacaa cgtctatatc accgccgaca
     2221 agcagaagaa cggcatcaag gccaacttca agatccgcca caacgtggag gacggcagcg
     2281 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     2341 ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac gagaagcgcg
     2401 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactcacggc agctccggct
     2461 ccagcgggtc tagcatggat agcactgaga gcctgttcac cggggtggtg cccatcctgg
     2521 tcgagctgga cggcgacgta aacggccaca agttcagcgt gcgcggcgag ggcgagggcg
     2581 atgccaccaa cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     2641 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     2701 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     2761 gcaccatcac cttcaaggac gacggcacct acaagacccg cgccgaggtg aagttcgagg
     2821 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     2881 tcctggggca caagctggag tacaacttca acagccacaa cgtctatatc accgccgaca
     2941 agcagaagaa cggcatcaag gccaacttca agatccgcca caacgtggag gacggcagcg
     3001 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     3061 ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac gagaagcgcg
     3121 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactcacggc agctccggct
     3181 ccagcgggta aagcggccgc tcctctggcg tttcttatca atcctttctc ttctctatgt
     3241 atatttttag aaggataata cctctatgta gacaatctta taaagatctt cgcaacaata
     3301 catatttcac ttactaatat atgttgcgtt ttcatgcatc aaaagtagtt agcaatgcta
     3361 gatcgtgtaa aggaaaaaaa agagaaatag tataattcgc catcctcata accactgaaa
     3421 aatcgaagtt aaccaaaagg aaagttgaaa tggaggaagt gttccgtttt tactcgaact
     3481 ctagaaatat cttcattcat aaaagccttt cattaaaacc atccaccatt gatgacccca
     3541 aaagtggtac cgagctcgaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     3601 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     3661 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     3721 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc
     3781 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     3841 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     3901 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     3961 cgaaagggcg ggtaataact gatataatta aattgaagct ctaatttgtg agtttagtat
     4021 acatgcattt acttataata cagtttttta gttttgctgg ccgcatcttc tcaaatatgc
     4081 ttcccagcct gcttttctgt aacgttcacc ctctacctta gcatcccttc cctttgcaaa
     4141 tagtcctctt ccaacaataa taatgtcaga tcctgtagag accacatcat ccacggttct
     4201 atactgttga cccaatgcgt ctcccttgtc atctaaaccc acaccgggtg tcataatcaa
     4261 ccaatcgtaa ccttcatctc ttccacccat gtctctttga gcaataaagc cgataacaaa
     4321 atctttgtcg ctcttcgcaa tgtcaacagt acccttagta tattctccag tagataggga
     4381 gcccttgcat gacaattctg ctaacatcaa aaggcctcta ggttcctttg ttacttcttc
     4441 tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc acaccgtgtg cattcgtaat
     4501 gtctgcccat tctgctattc tgtatacacc cgcagagtac tgcaatttga ctgtattacc
     4561 aatgtcagca aattttctgt cttcgaagag taaaaaattg tacttggcgg ataatgcctt
     4621 tagcggctta actgtgccct ccatggaaaa atcagtcaag atatccacat gtgtttttag
     4681 taaacaaatt ttgggaccta atgcttcaac taactccagt aattccttgg tggtacgaac
     4741 atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg atattaaata gcttggcagc
     4801 aacaggacta ggatgagtag cagcacgttc cttatatgta gctttcgaca tgatttatct
     4861 tcgtttcctg catgtttttg ttctgtgcag ttgggttaag aatactgggc aatttcatgt
     4921 ttcttcaaca ctacatatgc gtatatatac caatctaagt ctgtgctcct tccttcgttc
     4981 ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa agaaaccgaa atcaaaaaaa
     5041 agaataaaaa aaaaatgatg aattgaattg aaaagctggc ctcgtgatac gcctattttt
     5101 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     5161 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5221 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5281 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5341 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5401 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5461 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     5521 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5581 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5641 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5701 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5761 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5821 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5881 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5941 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6001 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6061 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6121 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6181 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6241 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6301 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6361 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6421 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     6481 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6541 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6601 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6661 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6721 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6781 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6841 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6901 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6961 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     7021 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag a
//
