LOCUS       sequence-420126-        5653 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pkk223 with Marinosulfonomonas MutY containing F302A, T303A, and 
            H304A mutations to reduce recognition of OG:A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5653)
  AUTHORS   Utzman PH, Mays VP, Miller BC, Fairbanks MC, Brazelton WJ, Horvath 
            MP
  TITLE     Metagenome mining and functional analysis reveal oxidized guanine 
            DNA repair at the Lost City Hydrothermal Field.
  JOURNAL   PLoS One. 2024 May 8;19(5):e0284642. doi: 
            10.1371/journal.pone.0284642. eCollection 2024.
  PUBMED    38718041
REFERENCE   2  (bases 1 to 5653)
  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: "PLoS One.";
            date: "2024-05-8"; volume: "19(5):e0284642. doi"; pages: " 
            10.1371/journal.pone.0284642. eCollection 2024"
COMMENT     Sequence Label: pkk223_MsMutY_F302A_T303A_H304A
FEATURES             Location/Qualifiers
     source          1..5653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1135..1152)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1305..1391
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1483..1510
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1529..1620
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1621..1689,1690..2481)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1839..1858)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2652..3240
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3141..3160
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3394..3411
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3426..3566
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3492..3511)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3652..3674
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3668..3859)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5317..5336
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(5583..5588,5589..5604,5605..5611)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5619..5635
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5624..5646
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
ORIGIN
        1 attcggtgac catgcgcgac taccgcgaca actccgaact ggggaccgtc cttctggggt
       61 ggtacgatgc taacgcgaga agcttaccct ggcgtgtagc cccagcacgt tcgaaagcag
      121 gggctttgcc ggatccatac cgtatttgga tgagcgaagt catgctgcag caaacgactg
      181 tcgctgcagt cgctaaatat tttcacgcgt ttacgactcg ctggccaacc attatcgcct
      241 tggcatctgc caatgacgat gatgttatgg gacagtgggc aggcctgggt tactatgctc
      301 gtgcacgcaa ccttttgaag tgtgcacgcg tggtagtagc ggattatggc ggcaatttcc
      361 ccgacacacg cgatgagttg cttaaacttc ctggcattgg accctatacg gcggcagcta
      421 ttgcaagtat cgccttcggt cgttcagaaa cagtgatgga tgcgaatatc gaacgcgtga
      481 tggcacgtat gcatgacatc cattcgccgt tgcccagtgc taagcctgaa cttcttgcat
      541 gcgcggcagc ccttaccccg gatttccgtg ccggcgatta tgctcaagct gtaatggact
      601 taggcgccac catttgcacg ccgaagtctg catcttgtgg agtgtgcccc tggtccttcg
      661 catgcctagc ccgtatcaat ggtacagctg ccgtcttacc gaaaaagact cctaaacccg
      721 tcaagccaac acgctacgga atcgcgtaca tcgcaagacg tggggacggt gcgtggttgt
      781 tagaacgccg tcctgataaa ggtttactgg gaggaatgct gggctggccc ggggctcagt
      841 ggggcgatca ggcggtagac aagccgcctg tttcagctga ttggcaagac cccggtgccg
      901 aagtgcgcca ttcagctgcg gcctttcatc tgcgtttatc gttacgcatt agcgtcgtcg
      961 gtaatgcgaa accaaccgca gggcacttca tcgcggatgc agattttgac cccgagacac
     1021 tacccaccgt aatgcgtaaa gcatatcgta ttgcacgcac aaagtttgac ggcgacgcga
     1081 acggttgact gcagccaagc ttggctgttt tggcggatga gagaagattt tcagcctgat
     1141 acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag
     1201 cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg
     1261 tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata aaacgaaagg
     1321 ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga
     1381 gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc
     1441 gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc atcctgacgg
     1501 atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca ttcaaatatg
     1561 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     1621 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     1681 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     1741 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     1801 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     1861 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     1921 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     1981 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2041 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2101 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2161 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2221 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2281 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     2341 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     2401 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     2461 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     2521 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     2581 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     2641 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     2701 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     2761 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     2821 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     2881 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     2941 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3001 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3061 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3121 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3181 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3241 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3301 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3361 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     3421 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatatgg
     3481 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     3541 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     3601 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     3661 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     3721 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     3781 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     3841 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
     3901 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     3961 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     4021 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     4081 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     4141 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     4201 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     4261 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     4321 cccgtggcca ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg gagatggcgg
     4381 acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc cgcaagaatt
     4441 gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg cttccattca
     4501 ggtcgaggtg gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga caaggtatag
     4561 ggcggcgcct acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg cataaatcgc
     4621 cgtgacgatc agcggtccaa tgatcgaagt taggctggta agagccgcga gcgatccttg
     4681 aagctgtccc tgatggtcgt catctacctg cctggacagc atggcctgca acgcgggcat
     4741 cccgatgccg ccggaagcga gaagaatcat aatggggaag gccatccagc ctcgcgtcgc
     4801 gaacgccagc aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt
     4861 ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat
     4921 tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc
     4981 gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt
     5041 cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     5101 ggctctcaag ggcatcggtc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc
     5161 cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat
     5221 ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct
     5281 catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc
     5341 agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatccg
     5401 ggcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
     5461 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
     5521 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
     5581 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
     5641 cacaggaaac aga
//
