LOCUS       sequence-420120-        5689 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pkk223 with Thiotrichaceae MutY insert.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5689)
  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 5689)
  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_TtMutY
FEATURES             Location/Qualifiers
     source          1..5689
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1180..1197)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1350..1436
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1528..1555
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1574..1665
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1666..1734,1735..2526)
                     /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(1884..1903)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2697..3285
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3186..3205
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3439..3456
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3471..3611
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3537..3556)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3697..3719
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3713..3904)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5362..5381
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(5628..5633,5634..5649,5650..5656)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    5664..5680
                     /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     join(5669..5689,1..2)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
ORIGIN
        1 ggaaacagaa ttcggtgacc atgggtgcga gcaagtggcg cgtctttgaa gctgtcatcc
       61 aatttcccga gaagatccta gtgtggtttg acgtgcacgg gcgtaaggac cttccgtggc
      121 aaaaaaatat taatccgtac cgtgtttggg tgtcggagat tatgcttcag caaacccagg
      181 ttaagacagt tattccctac ttcgagaaat tcatggctac tttcccagac gttcaaactt
      241 tagctcacgc tgaccaggac ttagtattga aacactggtc cggactgggc tactacgctc
      301 gcgcgcgtaa catgcataaa gctgcaaaaa tgatctgcgg agactttaaa ggccagtttc
      361 ctgacaatct ggaggctatg caaagcttgt caggcatcgg gcgctcgacc gccgccgcta
      421 tcttgagtat tgcctcgaat cagccgcagg ctattctgga cggaaatgtg aagcgcgtct
      481 tgactcgtgt gttcgtggtg gatggttggc ctggaaaggc cgatgtcctt aaaaagttgt
      541 ggttattggc tgagcagaca acacctgcaa gccgcaacgc tgattataca caagcaatta
      601 tggaccttgg agcaacctta tgtactcgtt ctaagccgcg ttgtgaagac tgtcctctaa
      661 aagccgactg tttagcgttc caacagggga atcagacagt ctaccctact tccaaaccaa
      721 agaaagttat gcccgaaaaa caggccatca tggtaattct aaagaattct aaacaggaag
      781 tcttcatgca aaaacgcccg ccagtgggca tctggggatc tttgtggtgc ttcccccagt
      841 ttgaaaagcg cgcctttgct gaggagtggt tgaaatctaa ttatggtaag ttaattcagg
      901 aaagtattgc gcttgcagaa atcaaacaca ctttttcgca ttttcatctg tatatccagc
      961 cattaatcat taatgttgaa acaccaattg agttgggtgt catggaggac aacgaatctc
     1021 tttggtataa cattacgacg gagtttaatg gtggtcttgc ggctcccgtc caaactctaa
     1081 ttaacattac caagaacatc gagaaagaga ttgagaatgg aaccaacggc tgactgcagc
     1141 caagcttggc tgttttggcg gatgagagaa gattttcagc ctgatacaga ttaaatcaga
     1201 acgcagaagc ggtctgataa aacagaattt gcctggcggc agtagcgcgg tggtcccacc
     1261 tgaccccatg ccgaactcag aagtgaaacg ccgtagcgcc gatggtagtg tggggtctcc
     1321 ccatgcgaga gtagggaact gccaggcatc aaataaaacg aaaggctcag tcgaaagact
     1381 gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc
     1441 cgggagcgga tttgaacgtt gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc
     1501 cataaactgc caggcatcaa attaagcaga aggccatcct gacggatggc ctttttgcgt
     1561 ttctacaaac tcttttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     1621 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     1681 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     1741 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     1801 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     1861 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg
     1921 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     1981 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     2041 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     2101 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     2161 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     2221 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     2281 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     2341 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     2401 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     2461 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     2521 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     2581 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     2641 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     2701 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     2761 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     2821 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     2881 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2941 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     3001 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     3061 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     3121 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     3181 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     3241 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     3301 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     3361 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     3421 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     3481 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tatggtgcac tctcagtaca
     3541 atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg
     3601 tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     3661 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     3721 tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt
     3781 gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa
     3841 gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg
     3901 tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata ccgatgaaac
     3961 gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta ctggaacgtt
     4021 gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc actcagggtc
     4081 aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag cagcatcctg
     4141 cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc agactttacg
     4201 aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc
     4261 agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta aggcaacccc
     4321 gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt ggccaggacc
     4381 caacgctgcc cgagatgcgc cgcgtgcggc tgctggagat ggcggacgcg atggatatgt
     4441 tctgccaagg gttggtttgc gcattcacag ttctccgcaa gaattgattg gctccaattc
     4501 ttggagtggt gaatccgtta gcgaggtgcc gccggcttcc attcaggtcg aggtggcccg
     4561 gctccatgca ccgcgacgca acgcggggag gcagacaagg tatagggcgg cgcctacaat
     4621 ccatgccaac ccgttccatg tgctcgccga ggcggcataa atcgccgtga cgatcagcgg
     4681 tccaatgatc gaagttaggc tggtaagagc cgcgagcgat ccttgaagct gtccctgatg
     4741 gtcgtcatct acctgcctgg acagcatggc ctgcaacgcg ggcatcccga tgccgccgga
     4801 agcgagaaga atcataatgg ggaaggccat ccagcctcgc gtcgcgaacg ccagcaagac
     4861 gtagcccagc gcgtcggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
     4921 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
     4981 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
     5041 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     5101 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
     5161 cggtcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     5221 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     5281 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     5341 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     5401 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atccgggctt atcgactgca
     5461 cggtgcacca atgcttctgg cgtcaggcag ccatcggaag ctgtggtatg gctgtgcagg
     5521 tcgtaaatca ctgcataatt cgtgtcgctc aaggcgcact cccgttctgg ataatgtttt
     5581 ttgcgccgac atcataacgg ttctggcaaa tattctgaaa tgagctgttg acaattaatc
     5641 atcggctcgt ataatgtgtg gaattgtgag cggataacaa tttcacaca
//
