LOCUS       sequence-424626-        7400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integrates onto p1 a partial LEU2 sequence to be subsequently used 
            as an integration flank.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7400)
  TITLE     Continuous evolution of user-defined genes at 1-million-times the 
            genomic mutation rate
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7400)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pGR420
FEATURES             Location/Qualifiers
     source          1..7400
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1539..1561
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1599..1617)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1685..1789
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1790..1858,1859..2650)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEVDKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2008..2027)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2821..3409
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3310..3329
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3563..3580
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             4051..5385
                     /codon_start=1
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /product="O-acetylserine and O-acetylhomoserine 
                     sulfhydrylase, required for sulfur amino acid synthesis"
                     /label=MET17
                     /translation="MPSHFDTVQLHAGQENPGDNAHRSRAVPIYATTSYVFENSKHGSQ
                     LFGLEVPGYVYSRFQNPTSNVLEERIAALEGGAAALAVSSGQAAQTLAIQGLAHTGDNI
                     VSTSYLYGGTYNQFKISFKRFGIEARFVEGDNPEEFEKVFDERTKAVYLETIGNPKYNV
                     PDFEKIVAIAHKHGIPVVVDNTFGAGGYFCQPIKYGADIVTHSATKWIGGHGTTIGGII
                     VDSGKFPWKDYPEKFPQFSQPAEGYHGTIYNEAYGNLAYIVHVRTELLRDLGPLMNPFA
                     SFLLLQGVETLSLRAERHGENALKLAKWLEQSPYVSWVSYPGLASHSHHENAKKYLSNG
                     FGGVLSFGVKDLPNADKETDPFKLSGAQVVDNLKLASNLANVGDAKTLVIAPYFTTHKQ
                     LNDKEKLASGVTKDLIRVSVGIEFIDDIIADFQQSFETVFAGQKP"
ORIGIN
        1 caatattccc acaagcattc cttctcaact catacgagct ttgttcagcc actgcccagc
       61 ccctgggccc ctcgctgacg catggaagaa ccctttcctt ttccttttcc ctgatctata
      121 aattgcagcc tcacaatacg taattgcaac attacaagcc ttatttttca aagaagatca
      181 tttaaacaaa tttttaaagg tttttttccc atatataaca acaacttgaa acagtttaga
      241 ggcttgtgtc taccagttca gttgctgcat tacgtttttt ggctagatta gcatcgctgg
      301 ttgatgctat taggttgtgc ataaacttaa acgtgttttt ttgggtctga tttcaagttc
      361 tcgcttttag tttgcttctg ttctaaactt ttaaagggaa aacaatatcg tattttccgg
      421 gaaaacccat cttacttaga gaaagggaag gaaaatacta ggatttacac tagcgattgc
      481 agtttcgata gttgcaattc tgatattctg gcattattct gcttaatcaa caagtgtttt
      541 gcgtttattt tatctaataa gatcaataag cgatttctaa atcaaagggt aaaatatcat
      601 atcaaagagg gttttttaaa aaacatataa gcaaggcacc ctttcgtacg ttgacaacac
      661 ttagggaagc gataggttta acgatcattt tgatatcaga tttctagagt tttaaacgtt
      721 tccttgttgt tttatttcat atttgatatc ttcatttgct gatttaatcc ataatggctt
      781 catctaatgt ggactcttct gagccccgta tttccagaga aagctctttg aagaggtctc
      841 tatctatttc aaagtcgtta aaagggttgt tcaagtcagg agggggaaac gccaatactg
      901 gccctactac tgctgctgct gctgctgcac catctagtat ccatgagcca ccattgccta
      961 tttggtcctt ggataaagct aatgttttgg cctcttcaag attatggaga aaaactgtgg
     1021 aggaaaccat caagaacgaa ttccctacat tgaaggttca acatcaattg attgattctg
     1081 ccgccatgat cctagttaag aacccaaccc acctaaatgg tattataatc accagcaaca
     1141 tgtttggtga tatcatctcc gatgaagcct ccgttatccc aggttccttg ggtttgttgc
     1201 catctgcgtc cttggcctct ttgccagaca agaacaccgc atttggtttg tacgaaccat
     1261 gccacggttc tgctccagat ttgccaaaga ataaggtcaa ccctatcgcc actatcttgt
     1321 ctgctgcaat gatgttgaaa ttgtcattga acttgcctga agaaggtaag gccattgaag
     1381 atgcagttaa aaaggttttg gatgcaggta tcagaactgg tgatttaggt ggttccaaca
     1441 gtaccaccga agtcggtgat gctgtcgccg aagaagttaa gaaaatcctt gcttaatttt
     1501 aaattaataa tgtaattcat atattatagt actcgtctcc gggagctgca tgtgtcagag
     1561 gttttcaccg tcatcaccga aacgcgtgag acgaaagggc ctcgtgatac gcctattttt
     1621 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     1681 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1741 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1801 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     1861 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     1921 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     1981 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     2041 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     2101 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     2161 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     2221 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     2281 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     2341 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     2401 attaatagac tggatggagg tggataaagt tgcaggacca cttctgcgct cggcccttcc
     2461 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2521 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2581 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2641 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     2701 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2761 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2821 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2881 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2941 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     3001 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     3061 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     3121 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3181 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3241 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3301 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3361 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3421 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3481 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3541 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     3601 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     3661 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat agtactatct attttttcag
     3721 tacataattt aattaaatca tcatgacctt tatctaattt tacatctctt ctaccaagac
     3781 ctaattcttt attaatacta tcaaaatctg ttattctatt gccttgagga tctttgaaac
     3841 accaatctga gtctgaggga tcttgtatag ataaaaaatt acgtatatca tttatagatg
     3901 gagaagttaa taaattttct aaaagaggaa aattaatttc taatgtgaac actagtgaga
     3961 tagctaaaga tcttaattgt gaaaacaata ttgaaagtat aataaataca ttaaaagaac
     4021 aaaatagata ttttgacaaa caaattgcat atgccatctc atttcgatac tgttcaacta
     4081 cacgccggcc aagagaaccc tggtgacaat gctcacagat ccagagctgt accaatttac
     4141 gccaccactt cttatgtttt cgaaaactct aagcatggtt cgcaattgtt tggtctagaa
     4201 gttccaggtt acgtctattc ccgtttccaa aacccaacca gtaatgtttt ggaagaaaga
     4261 attgctgctt tagaaggtgg tgctgctgct ttggctgttt cctccggtca agccgctcaa
     4321 acccttgcca tccaaggttt ggcacacact ggtgacaaca tcgtttccac ttcttactta
     4381 tacggtggta cttataacca gttcaaaatc tcgttcaaaa gatttggtat cgaggctaga
     4441 tttgttgaag gtgacaatcc agaagaattc gaaaaggtct ttgatgaaag aaccaaggct
     4501 gtttatttgg aaaccattgg taatccaaag tacaatgttc cggattttga aaaaattgtt
     4561 gcaattgctc acaaacacgg tattccagtt gtcgttgaca acacatttgg tgccggtggt
     4621 tacttctgtc agccaattaa atacggtgct gatattgtaa cacattctgc taccaaatgg
     4681 attggtggtc atggtactac tatcggtggt attattgttg actctggtaa gttcccatgg
     4741 aaggactacc cagaaaagtt ccctcaattc tctcaacctg ccgaaggata tcacggtact
     4801 atctacaatg aagcctacgg taacttggca tacatcgttc atgttagaac tgaactatta
     4861 agagatttgg gtccattgat gaacccattt gcctctttct tgctactaca aggtgttgaa
     4921 acattatctt tgagagctga aagacacggt gaaaatgcat tgaagttagc caaatggtta
     4981 gaacaatccc catacgtatc ttgggtttca taccctggtt tagcatctca ttctcatcat
     5041 gaaaatgcta agaagtatct atctaacggt ttcggtggtg tcttatcttt cggtgtaaaa
     5101 gacttaccaa atgccgacaa ggaaactgac ccattcaaac tttctggtgc tcaagttgtt
     5161 gacaatttaa agcttgcctc taacttggcc aatgttggtg atgccaagac cttagtcatt
     5221 gctccatact tcactaccca caaacaatta aatgacaaag aaaagttggc atctggtgtt
     5281 accaaggact taattcgtgt ctctgttggt atcgaattta ttgatgacat tattgcagac
     5341 ttccaacaat cttttgaaac tgttttcgct ggccaaaaac catgacatac cggatcattc
     5401 gttgaatttt ttgcatattt ataagtacag tttgttgaaa cagtacgttt tcgttccaat
     5461 cggtgtgata ttgatcatga tgatcttata tggagttaat atattaatta aagatgtgat
     5521 agggatcaaa tttcctgcat cagttgctgt aatgttggta aatttctgtt tcttatgtgt
     5581 tttatctatg attaatgaga acttaactca gaagattgtt ggtgtcatcg atgttccttt
     5641 aagttgggct ttgagatgga tgaatttatt ctttacacca gcgtttgtca ctttaccttt
     5701 gagtccctgg atatctttta gagaagctat gttaatagct gcctcgttcg tgttcatgta
     5761 cataatcacc gggttttgtt tggcatacgt aactatattt ggtcaaaagc tgtttgcact
     5821 ggctactggc caccttggta ggtcgaatgg aaggaaacgg acttctttac tcggcagtaa
     5881 agaggatatt cttcgtgatg ctgagaatgg atacagttct gcggatgaag atgctactga
     5941 agaagaaagt acactggaca taaacgaaga cgatgatgat attcatagtt ttgctgacat
     6001 tttctcggat cctgatgttg aagatttcca attagtcaac gttgcttcaa atagtgggtt
     6061 cccattgcaa agaactcgta caaatcttac tcatcatttg gcaatgggtg atactgctaa
     6121 tttagatgct gacattctaa gaggtagtga aattggactt cctaacgaca agggattagg
     6181 gatcactcaa aatgtgcagg acatttatga cgaggataac actcatggat ctgactcaac
     6241 tgctgaaaca aatcagttgt tattacagcc tgctctgata cattctcgta ataatgcagc
     6301 catggttcca atggcatcga aacgtttctg tggatctacc tctcatgatc aattttctga
     6361 tagaaataga gaggtagagt gtgaaagagc catcaccagg cagttctctc acagaatgga
     6421 acatcttttc accgtaaaca tgtggcacga tcacttgcat catgttctat acggattggg
     6481 gtttttcgca actatattca catactattt ttcatggtac atcatgcctt tccaactatt
     6541 taccgccatt tgcatgttct tcatcgtcac cgatgcacca ttattgccaa atcctaagta
     6601 taagaaattc ttgcatcctg ttatttgttc agtggctcta ttctggatcg ttgaattgat
     6661 ctcatcactc attaaacatc gccaaatcaa atatttcctc tcggatcttc gtgaatacaa
     6721 ggttggtaca acatacttaa cattattcac aaatactagt gtatcaggtc caaacaagaa
     6781 ctggcctggc gccggtgata ttttttcctc ctgtatggac gtctctattg tcgcattatc
     6841 gatgcccatg tacacttaca gaagcgatct aaagaaacat tttgtcgtgc ttcttatccc
     6901 aaccatctct ttatgtgctg ccactctaat gttaaatccc tttatatgct acaacatcgg
     6961 tatcagctca tataactcca ttggattcat tggaagaagc atcactttag ccttggggat
     7021 gcccgtcatc gaaaatttcg aaggatccgt taccgtaatg gctgtcatta cagtgctaag
     7081 tggtatcgta ggtgcactat ccggaggtcc catgcttgac ttcttcagag tacctcgcga
     7141 tgatttcgtc accagaggcc taactctagg ctgtaactgc ggtgccatcg ccaccagcta
     7201 cctcttaaca gtagatagaa gagctgctgc tatcagctcc ctttccttcg tcttatacgg
     7261 tgcaataatg gttatcctaa gttcgatagg gcccataaaa gacttcgtgc atcagatcgt
     7321 atcgctataa ccaaatgatc caactactga tagcccactt tacacaattc tctcacatat
     7381 acaacacata tacacatatt
//
