LOCUS       sequence-375962-        7389 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli - Eggerthella lenta shuttle plasmid (KanR), cumate-inducible
            promoter Pct5.1-crRNA(dadR-targeting spacer).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7389)
  AUTHORS   Dong X, Guthrie BGH, Alexander M, Noecker C, Ramirez L, Glasser NR, 
            Turnbaugh PJ, Balskus EP
  TITLE     Genetic manipulation of the human gut bacterium Eggerthella lenta 
            reveals a widespread family of transcriptional regulators.
  JOURNAL   Nat Commun. 2022 Dec 9;13(1):7624. doi: 10.1038/s41467-022-33576-3.
  PUBMED    36494336
REFERENCE   2  (bases 1 to 7389)
  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: "Nat 
            Commun."; date: "2022-12-9"; volume: "13(1):7624. doi"; pages: " 
            10.1038/s41467-022-33576-3"
COMMENT     Sequence Label: pXD71Cas10.1-Pct5.1-crRNA(dadR)
FEATURES             Location/Qualifiers
     source          1..7389
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3013..3807
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     terminator      complement(3930..3957)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    4090..4117
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     protein_bind    4132..4159
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     CDS             4497..5108
                     /codon_start=1
                     /gene="cymR from Pseudomonas putida"
                     /product="cumate repressor (Mullick et al., 2006)"
                     /label=CymR
                     /translation="MSPKRRTQAERAMETQGKLIAAALGVLREKGYAGFRIADVPGAAG
                     VSRGAQSHHFPTKLELLLATFEWLYEQITERSRARLAKLKPEDDVIQQMLDDAAEFFLD
                     DDFSISLDLIVAADRDPALREGIQRTVERNRFVVEDMWLGVLVSRGLSRDDAEDILWLI
                     FNSVRGLAVRSLWQKDKERFERVRNSTLEIARERYAKFKR"
     misc_feature    5111..5130
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that start with ""ATG"""
     primer_bind     complement(5287..5305)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5373..5477
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5478..5546,5547..6338)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5696..5715)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6509..7097
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6998..7017
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(7278..7349)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttcgagacaa ggagattata gcatggcggc aaacgctggc gagacgaagg tgaagctctg
       61 ggagggcatc tgctacccag agaacatgcg cgatgactgg caggacgaga tagacgacat
      121 tcttcaagtg cctttcgctt atgcggttca cgacatagac catgaccaga agagcaagca
      181 gcgcaagacg catgcgcacg tcattgttgt ctggggcggc aacacgacac gcaaggcaat
      241 catcaacgtc ttgaaccgat tgagcgcgga cggcaagaag tgctgttcgt cagccgagcc
      301 aatcaacaac atcaggcatg cgtatgacta tctgattcac gacacggctt cctgtcgcaa
      361 gaagggcaag gagctgtatc cggtcgaagc acgcatcgag ggaaacaact tcgacattgg
      421 tcagctcgaa cagctctcga ctaaggacaa gcaggacatg ctcttcgagt tggtcggctt
      481 catcatgtgc gagaggttcg agaccatcaa cgatttcacg gctgcggcgc tgagggagtt
      541 tcccgagcag taccgcgaaa tcatcgtggg gtacaactcg attctcgagc gctattgtcg
      601 cgggaactat ctgaacgccg aacgtaagcg caagtgcgtg gaacagtcag gtgccaaaga
      661 gtagtcgggt caatgcagca atgcgtgact acggggcggc gtggtctgcg ctcgcgcaga
      721 tctaagccga ctcgttcacg gattgcggaa ttgacgcgac ggccaaccga atagaccaga
      781 cttgattaag ggtcgatgga cgctctgctc catcgaccct ttccttgccc gtgtctcgac
      841 tggatgagac ctgcagaatc gggcggctca ttatttttcg ctaactacgt ccgaaaaata
      901 atgtgtccga aaaataatgt gacaaatgac aaatgacaaa tgaccacagg ggggccacta
      961 cgacggcccc cctaatagtg ccgtgtgccg tgtgccgaac cgcaggtaga ttgaggggtg
     1021 gttcggaggg gcgaagtccc tccgttcggc gctcgcgccg atggcccgag ggccgtgagc
     1081 gaagccgcga acgggcaagg gctggggggt ctcgggggga acgcccgagc gggtgcaggg
     1141 ctggccctgc cacgtcctta gcgtagctaa ggtctaacgt gctagacttt agtcgattaa
     1201 gtctgtacgg ttcggaggtt tccatggctc acatcgcgaa atacaaggct ccatcggtcg
     1261 ggcacatgct tgcccactac cgccgcgacc gcagcagcct ggagcgcgac aacatcgacc
     1321 ccgagcgcat caagaacgac ttggtggtcg gccactacac gaacaaggac ggcaagctcg
     1381 ttgtggggcg cgtggagccg cgcgatggcg agccgaactg gggcacggtg gagagccgca
     1441 tcgagcgcgt gaacgaggcc cagaaggccg cagggaagcg tgccacgcgc aaggatgcgg
     1501 tcgttatggc tgacgttgtt gtgacgcttc ccgacaatgt gcgcaagggc gatgaggaca
     1561 ggttcttccg tctcacgtac tggtacctgt cgaagaagtt cggcatcgag aacatgatgg
     1621 gcgggtacgt ccacaaggac gaggtgttga aggatggcac gcccgcccgc gaccacatgc
     1681 acgtgccgtt cacgcccatc ctagatggac ggttcaacta caagcagatg tgcccccgca
     1741 gcttctaccg gagcatgcac aaggagcttg gggattacct agaggggcgc atgggctacc
     1801 gccctgcaat cgagcttgac gaggagacgc gggcgcagcg tgtatacacg gacaagtccg
     1861 tggatatcga caaggtacgc tgtgcggtcg ataaggccgt ggttgaacct gcgaaggacg
     1921 aagcggcgcg aatcgtcgcg gcggctaggg aagaagccgc cgctttactg aatgatgccg
     1981 aaacgcgtaa ggcagagctt gttaccgaga ttgccgacaa ggagggtgac cttgccgagc
     2041 ttgacagcca gctcgaggac gtgaagctcg atatcgaaga tgagcaagac cgattggagt
     2101 gtcttcggca acgagcggac ggagttgcgc gagacattgg agagcttcaa cccatcgcta
     2161 ccgaggttcg gggctgggaa gctgcgggaa aagctgagcg cggcgcaatc ctcgataaca
     2221 tcgttgttaa gtgcgatgga ctcgcgagcc gcatcagagc aggagttgcg ggaatgcgaa
     2281 tcaaagtgga agagcttaga agtcgaatat cgcgcccgct cgaatatctg atgcgcaggg
     2341 aacagccaag gcaacagagc cttaatgacg tgctgcgcga tgcgcagcgg gcggcggatg
     2401 cgtggaaccg cgaccacgct gcaccgcaga ggacgtatcg cgggcgggct agatgatgtg
     2461 ttatagtagt acctgtaata cacgacaagg aggtgcgacg cgaaaaaacc ccgctgaagc
     2521 ggggtttttt gcggagcttc gcttccacga tgcgcatgat tcggaggccc cggcgatcgc
     2581 cggggcctcc gtgcgttgtg tgtcgatttg ggatgcagcc agatgggctt cctcggcgcg
     2641 aaccggccga gatttccgaa tgcagacggt ctcgcttgga cgatgcgctt tcgaataatc
     2701 ccggcggagc gggcattcca gtcttcttat caatatatct atagaatggg caaagcataa
     2761 aaacttgcat ggactaatgc ttgaaaccca ggacaataac cttatagctt gtaaattcta
     2821 tcataattgt ggtttcaaaa tcggctccgt cgatactatg ttatacgcca actttgaaaa
     2881 caactttgaa aaagctgttt tctggtattt aaggttttag aatgcaagga acagtgaatt
     2941 ggagttcgtc ttgttataat tagcttcttg gggtatcttt aaatactgta gaaaagagga
     3001 aggaaataat aaatggctaa aatgagaata tcaccggaat tgaaaaaact gatcgaaaaa
     3061 taccgctgcg taaaagatac ggaaggaatg tctcctgcta aggtatataa gctggtggga
     3121 gaaaatgaaa acctatattt aaaaatgacg gacagccggt ataaagggac cacctatgat
     3181 gtggaacggg aaaaggacat gatgctatgg ctggaaggaa agctgcctgt tccaaaggtc
     3241 ctgcactttg aacggcatga tggctggagc aatctgctca tgagtgaggc cgatggcgtc
     3301 ctttgctcgg aagagtatga agatgaacaa agccctgaaa agattatcga gctgtatgcg
     3361 gagtgcatca ggctctttca ctccatcgac atatcggatt gtccctatac gaatagctta
     3421 gacagccgct tagccgaatt ggattactta ctgaataacg atctggccga tgtggattgc
     3481 gaaaactggg aagaagacac tccatttaaa gatccgcgcg agctgtatga ttttttaaag
     3541 acggaaaagc ccgaagagga acttgtcttt tcccacggcg acctgggaga cagcaacatc
     3601 tttgtgaaag atggcaaagt aagtggcttt attgatcttg ggagaagcgg cagggcggac
     3661 aagtggtatg acattgcctt ctgcgtccgg tcgatcaggg aggatatcgg ggaagaacag
     3721 tatgtcgagc tattttttga cttactgggg atcaagcctg attgggagaa aataaaatat
     3781 tatattttac tggatgaatt gttttagatc gttaagggat caactttggg agagagttca
     3841 aaattgatcc tttttttata acaggaatta aataataaaa aagccggatt aataatctgg
     3901 ctttttatat tctctctcta gtatataaac gcagaaaggc ccacccgaag gtgagccagt
     3961 gtgagtcgac tagtatttca acccgcactc cccatgcggg gagtgaccgc tcacccgctc
     4021 gcgccgcacg atgacggaga catttcaacc cgcactcccc atgcggggag tgacattagt
     4081 taatcataat acaaacagac cagattgtct gtttgttggc cgcaacactc gtacaaacag
     4141 accagattgt ctgtttgtta ctacatttat tgtacaacac gagcccattt ttgtcaaata
     4201 aaatttaaat tatatcaacg ttaataagac gttgtcaata aaattatttt gacaaaatag
     4261 aatagtaata caggatccga atcgtttcag ttgcctaaat aataaaaaag ccggattaat
     4321 aatcaaataa ccgttccttc aatgtttttt ttagaaaatt tagttataat attagatatg
     4381 atacttttaa atatctaatt caagcttcaa aaaacaccaa cttagttcgg tggataaaca
     4441 aaggagtggt tattattcaa attgcagatc aagctttagt aaaaaaaatg gtgatcatga
     4501 gtccaaagag aagaacacag gcagagcgcg caatggagac ccagggcaag ttgattgcag
     4561 cggccctggg ggttttacgg gaaaaaggtt acgcgggatt ccggatcgca gatgtgcccg
     4621 gtgctgctgg tgtctccaga ggagcgcaga gccatcattt cccgacaaag cttgagcttc
     4681 tgcttgccac ttttgaatgg ctttacgaac agatcaccga acgcagtcgg gctcgattag
     4741 cgaaattgaa gccagaggat gacgtcatcc agcaaatgct ggacgacgcc gccgaatttt
     4801 tcctcgacga tgacttctct atcagccttg atttgattgt ggctgccgac cgggacccag
     4861 cgttacgcga gggtattcag cgcacggtag agaggaatcg gtttgtcgtc gaggatatgt
     4921 ggcttggtgt tctggtgagc cgtggtcttt cgcgtgatga tgcagaagat atcctttggt
     4981 tgatattcaa ttcggtgcgt gggcttgctg ttcgtagcct atggcagaag gacaaagaac
     5041 gctttgagcg tgtcaggaac tcgacactcg aaattgcgcg agagcggtac gcgaaattca
     5101 agcgctagat ctagaagcgg ccgcgaattc agaatagtaa tacaggatcc gaatcgtttc
     5161 agttgccgcc ggattaataa tcaaataacc gttcctaaaa aatgccctct tgggttatca
     5221 agagggtcat tatatttcgc aaaaaacccc gcttcggcgg ggttttttcg ccttgaagac
     5281 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     5341 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     5401 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     5461 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5521 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     5581 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     5641 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     5701 gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact
     5761 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     5821 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     5881 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     5941 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     6001 agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     6061 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     6121 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     6181 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     6241 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     6301 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     6361 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     6421 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     6481 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     6541 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     6601 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     6661 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6721 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6781 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     6841 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6901 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     6961 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     7021 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     7081 ggcggagcct atggaaaaac gccagcaacg cggcccgaga tgcgccgcgt gcggctgctg
     7141 gagatggcgg acgcgatgga tatgttctgc caagggttgg tttgcgcatt cacagttctc
     7201 cgcaagaatt gattggctcc aattcttgga gtggtgaatc cgttagcgag gtgccgccgg
     7261 catgagctcc tctagtagag agcgttcacc gacaaacaac agataaaacg aaaggcccag
     7321 tctttcgact gagcctttcg ttttatttga aaaagaaaag ccccagcaag cttggcggca
     7381 gcaggggca
//
