LOCUS       Exported                7470 bp ds-DNA     circular SYN 07-JAN-2025
DEFINITION  Plasmid for tagging Sys1 with msGFP2x3 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7470)
  AUTHORS   Glick Lab
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Oct 2, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Linearize with BglII for integration at SYS1.
FEATURES             Location/Qualifiers
     source          1..7470
                     /organism="synthetic DNA construct"
                     /lab_host="Saccharomyces cerevisiae"
                     /mol_type="other DNA"
     rep_origin      complement(183..771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number colE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(942..1802)
                     /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"
     promoter        complement(1803..1907)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2005..2220
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             2221..3024
                     /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"
     primer_bind     3492..3508
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3516..3943
                     /label=5' SYS1
     CDS             3944..4552
                     /codon_start=1
                     /label=SYS1
                     /translation="MVSIRRYLRVPNELKPSQIFKQDSLSPSKIGLQIVLLQIFYYTTA
                     IVLFYCWAKLAGYDLNIKEWLFSWENIDFTNAYGLSISLLWLLDSLICVFFLTVIVGRS
                     KLAWDFAITIHAINFIVVFLYTRKFPSFSWFFLQILSSLILIFLGTWTTRWRELRDTFF
                     EGLVDPNEGEVGLVTPSQQHSNHSELEQSPIQLKDLESQI"
     CDS             4568..5275
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /note="yeast codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITGGSGGS"
     CDS             5282..5989
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /note="yeast codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITGGSGGS"
     CDS             5996..6703
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /note="yeast codon-optimized"
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITGGSGGS"
     misc_feature    6721..7232
                     /label=3' SYS1
     primer_bind     complement(7251..7267)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
       61 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      121 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
      181 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
      241 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
      301 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
      361 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
      421 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
      481 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
      541 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
      601 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
      661 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
      721 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
      781 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
      841 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
      901 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
      961 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1021 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1081 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1141 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     1201 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     1261 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     1321 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     1381 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     1441 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     1501 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     1561 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     1621 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     1681 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     1741 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     1801 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     1861 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     1921 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     1981 cgtatcacga ggccagcttt tcaattcaat tcatcatttt ttttttattc ttttttttga
     2041 tttcggtttc tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg
     2101 aaggagcaca gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat
     2161 tgcccagtat tcttaaccca actgcacaga acaaaaacat gcaggaaacg aagataaatc
     2221 atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag
     2281 ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc
     2341 accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca
     2401 catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta
     2461 tccgccaagt acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca
     2521 gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat
     2581 gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta
     2641 acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct
     2701 actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc
     2761 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg
     2821 acacccggtg tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg
     2881 gatgatgtgg tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag
     2941 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg
     3001 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac
     3061 tcacaaatta gagcttcaat ttaattatat cagttattac ccgccctttc gtctcgcgcg
     3121 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     3181 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     3241 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     3301 gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc
     3361 attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca
     3421 gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca
     3481 gtcacgacgt tgtaaaacga cggccagtga attcctggta atatttcagt tgttgactaa
     3541 tatatatagt ggtttttgcc aaaattaaag acttcctttt attaactagg tgaccagata
     3601 taaggaagtg tatttgcttt ctaataactt gtgtacggcc ataacccggc ttttaatcag
     3661 tttatttaac aaggtataca aatcgccaaa cgtaacaaga gataggcgct attattcatt
     3721 taatattgtc gtattcaaaa tactaacggt gaacaatttg ctccattgta acactatagg
     3781 acaaggctaa tcatgaataa atatctcccc acccaaaata ttcgtagctc gcgatattca
     3841 cggtaatatt ctgtaacttc atgttaagag atgatcttta ggaaggctaa ggaagggtac
     3901 acatagaagt gagataaaaa ataggcattt gaattactgt aacatggttt cgataagaag
     3961 gtatttgcga gtgccgaatg agttgaaacc ttcccagatc ttcaaacaag attctctctc
     4021 tccaagtaag attggacttc aaattgttct tttacagatt ttttattaca ccacagcgat
     4081 cgttcttttt tattgttggg cgaagctagc gggatatgat ctcaatatca aggaatggct
     4141 gttttcttgg gagaatatcg actttacgaa cgcatacggt ttatcaatct cactgttatg
     4201 gcttctggat tcattaatat gtgttttttt cttaactgtt atcgttggac ggagtaaact
     4261 cgcatgggat tttgctatca ctatacatgc catcaatttt attgtggtgt tcctctatac
     4321 tcggaaattt ccttcttttt cttggttttt tcttcaaatt ttatcatctt taatactaat
     4381 atttttgggg acatggacaa caagatggag agagcttaga gacacctttt ttgagggctt
     4441 ggttgatcct aatgaaggag aagttggatt ggttacgcct agtcagcagc atagtaatca
     4501 ttcagaattg gagcaatcac caatacaact aaaagactta gaaagccaaa tacgggatcc
     4561 aactagtatg gatagcactg agagcttgtt cacaggtgtt gttcctatct tggtcgaatt
     4621 agacggtgac gtaaacggtc ataagttcag tgttagaggt gaaggtgaag gtgacgctac
     4681 taacggtaaa ttgacattga agtttatttg tactacaggt aaattgccag ttccttggcc
     4741 aactttggtt actacattga catacggtgt tcaatgtttt tcaagatacc cagatcatat
     4801 gaagcaacat gatttcttta agtctgctat gccagaaggt tacgttcaag aaagaactat
     4861 cacttttaaa gatgatggta cttacaagac aagagcagag gttaagttcg aaggtgacac
     4921 tttggttaac agaatcgaat tgaagggtat cgattttaaa gaagatggta atattttggg
     4981 tcataaattg gaatacaact acaactcaca taacgtttac attacagctg ataaacaaaa
     5041 gaatggtatt aaagcaaact tcaagatcag acataatgtt gaagatggtt ctgttcaatt
     5101 agcagatcat taccaacaaa acactccaat tggtgacggt ccagttttgt taccagataa
     5161 ccattacttg tctacacaat caaaattgtc taaagatcca aacgaaaaga gagatcatat
     5221 ggttttgtta gaatttgtta ctgctgctgg tatcacaggt ggcagtggcg gttctgctag
     5281 tatggatagc actgagagct tgttcacagg tgttgttcct atcttggtcg aattagacgg
     5341 tgacgtaaac ggtcataagt tcagtgttag aggtgaaggt gaaggtgacg ctactaacgg
     5401 taaattgaca ttgaagttta tttgtactac aggtaaattg ccagttcctt ggccaacttt
     5461 ggttactaca ttgacatacg gtgttcaatg tttttcaaga tacccagatc atatgaagca
     5521 acatgatttc tttaagtctg ctatgccaga aggttacgtt caagaaagaa ctatcacttt
     5581 taaagatgat ggtacttaca agacaagagc agaggttaag ttcgaaggtg acactttggt
     5641 taacagaatc gaattgaagg gtatcgattt taaagaagat ggtaatattt tgggtcataa
     5701 attggaatac aactacaact cacataacgt ttacattaca gctgataaac aaaagaatgg
     5761 tattaaagca aacttcaaga tcagacataa tgttgaagat ggttctgttc aattagcaga
     5821 tcattaccaa caaaacactc caattggtga cggtccagtt ttgttaccag ataaccatta
     5881 cttgtctaca caatcaaaat tgtctaaaga tccaaacgaa aagagagatc atatggtttt
     5941 gttagaattt gttactgctg ctggtatcac aggtggcagt ggcggttctg ctagtatgga
     6001 tagcactgag agcttgttca caggtgttgt tcctatcttg gtcgaattag acggtgacgt
     6061 aaacggtcat aagttcagtg ttagaggtga aggtgaaggt gacgctacta acggtaaatt
     6121 gacattgaag tttatttgta ctacaggtaa attgccagtt ccttggccaa ctttggttac
     6181 tacattgaca tacggtgttc aatgtttttc aagataccca gatcatatga agcaacatga
     6241 tttctttaag tctgctatgc cagaaggtta cgttcaagaa agaactatca cttttaaaga
     6301 tgatggtact tacaagacaa gagcagaggt taagttcgaa ggtgacactt tggttaacag
     6361 aatcgaattg aagggtatcg attttaaaga agatggtaat attttgggtc ataaattgga
     6421 atacaactac aactcacata acgtttacat tacagctgat aaacaaaaga atggtattaa
     6481 agcaaacttc aagatcagac ataatgttga agatggttct gttcaattag cagatcatta
     6541 ccaacaaaac actccaattg gtgacggtcc agttttgtta ccagataacc attacttgtc
     6601 tacacaatca aaattgtcta aagatccaaa cgaaaagaga gatcatatgg ttttgttaga
     6661 atttgttact gctgctggta tcacaggtgg cagtggcggt tctgctagct aagcggccgc
     6721 gaggagatga aacaacatat tcgaggcaat tttggcaata ttatagtcat tcaagagaaa
     6781 acagtaaatg tatgaaaaat ggcaagaacc ttatctctat aggcttaata aatctgcctt
     6841 tttggccaat ttttcaaaac agatcctatt tcttatttat acacaatcta tatttaacat
     6901 aattaagtac gtgatgacat tgaagataat aataagtaaa gagtttacgt acttaaattt
     6961 tatatagttt tagtatactt gcaatgtgtg aatggatcta gtaagctgag gtacttctct
     7021 agcgtgaata tgaaacgagt aacagggtgg tacataattt acgaacagaa ctcaagttga
     7081 taaatccttt gcgttcttaa ccatttgtaa aagttcgtca acaacattga aaatattgct
     7141 tcgagcagat gttttatcgt ttctgcaaat ttccagtcct gaacacctct gttgtcgtaa
     7201 atgtaataat gccacactgg attcgattct agaagcttgg cgtaatcatg gtcatagctg
     7261 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     7321 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     7381 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     7441 gcggggagag gcggtttgcg tattgggcgc
//
