LOCUS       Exported                7122 bp ds-DNA     circular SYN 07-JAN-2025
DEFINITION  Plasmid for tagging Sys1 with Sys1-HaloTagx2 by gene replacement.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7122)
  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..7122
                     /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..5458
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /note="yeast codon-optimized"
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             5465..6355
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /note="yeast codon-optimized"
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             6367..6370
                     /codon_start=1
                     /gene="lacZ"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="G"
     misc_feature    6373..6884
                     /label=3' SYS1
     primer_bind     complement(6903..6919)
                     /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 gctgaaatcg gtacaggttt tccttttgac ccacactatg tcgaagtatt
     4621 aggtgaaaga atgcactatg ttgatgtcgg tcctagagat ggtactccag ttttgttttt
     4681 acatggtaac ccaacatctt catacgtttg gagaaacatc atcccacatg ttgcaccaac
     4741 tcatagatgt attgctccag atttgattgg tatgggtaaa tctgataagc cagatttggg
     4801 ttatttcttt gatgatcatg ttagattcat ggatgctttt attgaagcat tgggtttgga
     4861 agaagttgtt ttggttattc atgattgggg ttcagcatta ggttttcatt gggctaagag
     4921 aaacccagaa agagttaagg gtatcgcttt tatggaattc attagaccaa ttccaacttg
     4981 ggatgaatgg ccagaattcg caagagaaac attccaagct tttagaacta cagatgttgg
     5041 tagaaagttg atcatcgatc aaaacgtttt tattgaaggt actttgccaa tgggtgttgt
     5101 tagaccatta acagaagttg aaatggatca ttacagagaa ccatttttaa acccagttga
     5161 tagagaacca ttgtggagat ttccaaatga attaccaatt gctggtgaac cagcaaacat
     5221 cgttgctttg gttgaagaat acatggattg gttacatcaa tctccagttc caaagttgtt
     5281 attttggggt acaccaggtg ttttgattcc accagcagaa gctgcaagat tggctaagtc
     5341 attgccaaac tgtaaggctg ttgatattgg tccaggttta aatttgttac aagaagataa
     5401 tccagactta ataggttccg aaatcgccag atggttatca actttagaaa tcagtggtgc
     5461 tagtatggct gaaatcggta caggttttcc ttttgaccca cactatgtcg aagtattagg
     5521 tgaaagaatg cactatgttg atgtcggtcc tagagatggt actccagttt tgtttttaca
     5581 tggtaaccca acatcttcat acgtttggag aaacatcatc ccacatgttg caccaactca
     5641 tagatgtatt gctccagatt tgattggtat gggtaaatct gataagccag atttgggtta
     5701 tttctttgat gatcatgtta gattcatgga tgcttttatt gaagcattgg gtttggaaga
     5761 agttgttttg gttattcatg attggggttc agcattaggt tttcattggg ctaagagaaa
     5821 cccagaaaga gttaagggta tcgcttttat ggaattcatt agaccaattc caacttggga
     5881 tgaatggcca gaattcgcaa gagaaacatt ccaagctttt agaactacag atgttggtag
     5941 aaagttgatc atcgatcaaa acgtttttat tgaaggtact ttgccaatgg gtgttgttag
     6001 accattaaca gaagttgaaa tggatcatta cagagaacca tttttaaacc cagttgatag
     6061 agaaccattg tggagatttc caaatgaatt accaattgct ggtgaaccag caaacatcgt
     6121 tgctttggtt gaagaataca tggattggtt acatcaatct ccagttccaa agttgttatt
     6181 ttggggtaca ccaggtgttt tgattccacc agcagaagct gcaagattgg ctaagtcatt
     6241 gccaaactgt aaggctgttg atattggtcc aggtttaaat ttgttacaag aagataatcc
     6301 agacttaata ggttccgaaa tcgccagatg gttatcaact ttagaaatca gtggtgctag
     6361 ctaagcggcc gcgaggagat gaaacaacat attcgaggca attttggcaa tattatagtc
     6421 attcaagaga aaacagtaaa tgtatgaaaa atggcaagaa ccttatctct ataggcttaa
     6481 taaatctgcc tttttggcca atttttcaaa acagatccta tttcttattt atacacaatc
     6541 tatatttaac ataattaagt acgtgatgac attgaagata ataataagta aagagtttac
     6601 gtacttaaat tttatatagt tttagtatac ttgcaatgtg tgaatggatc tagtaagctg
     6661 aggtacttct ctagcgtgaa tatgaaacga gtaacagggt ggtacataat ttacgaacag
     6721 aactcaagtt gataaatcct ttgcgttctt aaccatttgt aaaagttcgt caacaacatt
     6781 gaaaatattg cttcgagcag atgttttatc gtttctgcaa atttccagtc ctgaacacct
     6841 ctgttgtcgt aaatgtaata atgccacact ggattcgatt ctagaagctt ggcgtaatca
     6901 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     6961 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     7021 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     7081 atcggccaac gcgcggggag aggcggtttg cgtattgggc gc
//
