LOCUS       sequence-375977-        8450 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli - Eggerthella lenta shuttle plasmid (KanR), cumate-inducible
            promoter Pct5.1-crRNA(cadR-targeting spacer) cadR-deleting repair 
            template,used for cadR deletion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8450)
  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 8450)
  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: pXD71Cas10.9RT2-Pct5.1-crRNA(cadR)-RT(Delta-cadR)
FEATURES             Location/Qualifiers
     source          1..8450
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3018..3812
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     terminator      complement(3935..3962)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    4095..4122
                     /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    4137..4164
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     CDS             4502..5113
                     /codon_start=1
                     /gene="cymR from Pseudomonas putida"
                     /product="cumate repressor (Mullick et al., 2006)"
                     /label=CymR
                     /translation="MSPKRRTQAERAMETQGKLIAAALGVLREKGYAGFRIADVPGAAG
                     VSRGAQSHHFPTKLELLLATFEWLYEQITERSRARLAKLKPEDDVIQQMLDDAAEFFLD
                     DDFSISLDLIVAADRDPALREGIQRTVERNRFVVEDMWLGVLVSRGLSRDDAEDILWLI
                     FNSVRGLAVRSLWQKDKERFERVRNSTLEIARERYAKFKR"
     misc_feature    5116..5135
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that start with ""ATG"""
     primer_bind     complement(5292..5310)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             6650..6661
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     rep_origin      7575..8163
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8064..8083
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(8344..8415)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gggcattcga gacaaggaga ttatagcatg gcggcaaacg ctggcgagac gaaggtgaag
       61 ctctgggagg gcatctgcta cccagagaac atgcgcgatg actggcagga cgagatagac
      121 gacattcttc aagtgccttt cgcttatgcg gttcacgaca tagaccatga ccagaagagc
      181 aagcagcgca agacgcatgc gcacgtcatt gttgtctggg gcggcaacac gacacgcaag
      241 gcaatcatca acgtcttgaa ccgattgagc gcggacggca agaagtgctg ttcgtcagcc
      301 gagccaatca acaacatcag gcatgcgtat gactatctga ttcacgacac ggcttcctgt
      361 cgcaagaagg gcaaggagct gtatccggtc gaagcacgca tcgagggaaa caacttcgac
      421 attggtcagc tcgaacagct ctcgactaag gacaagcagg acatgctctt cgagttggtc
      481 ggcttcatca tgtgcgagag gttcgagacc atcaacgatt tcacggctgc ggcgctgagg
      541 gagtttcccg agcagtaccg cgaaatcatc gtggggtaca actcgattct cgagcgctat
      601 tgtcgcggga actatctgaa cgccgaacgt aagcgcaagt gcgtggaaca gtcaggtgcc
      661 aaagagtagt cgggtcaatg cagcaatgcg tgactacggg gcggcgtggt ctgcgctcgc
      721 gcagatctaa gccgactcgt tcacggattg cggaattgac gcgacggcca accgaataga
      781 ccagacttga ttaagggtcg atggacgctc tgctccatcg accctttcct tgcccgtgtc
      841 tcgactggat gagacctgca gaatcgggcg gctcattatt tttcgctaac tacgtccgaa
      901 aaataatgtg tccgaaaaat aatgtgacaa atgacaaatg acaaatgacc acaggggggc
      961 cactacgacg gcccccctaa tagtgccgtg tgccgtgtgc cgaaccgcag gtagattgag
     1021 gggtggttcg gaggggcgaa gtccctccgt tcggcgctcg cgccgatggc ccgagggccg
     1081 tgagcgaagc cgcgaacggg caagggctgg ggggtctcgg ggggaacgcc cgagcgggtg
     1141 cagggctggc cctgccacgt ccttagcgta gctaaggtct aacgtgctag actttagtcg
     1201 attaagtctg tacggttcgg aggtttccat ggctcacatc gcgaaataca aggctccatc
     1261 ggtcgggcac atgcttgccc actaccgccg cgaccgcagc agcctggagc gcgacaacat
     1321 cgaccccgag cgcatcaaga acgacttggt ggtcggccac tacacgaaca aggacggcaa
     1381 gctcgttgtg gggcgcgtgg agccgcgcga tggcgagccg aactggggca cggtggagag
     1441 ccgcatcgag cgcgtgaacg aggcccagaa ggccgcaggg aagcgtgcca cgcgcaagga
     1501 tgcggtcgtt atggctgacg ttgttgtgac gcttcccgac aatgtgcgca agggcgatga
     1561 ggacaggttc ttccgtctca cgtactggta cctgtcgaag aagttcggca tcgagaacat
     1621 gatgggcggg tacgtccaca aggacgaggt gttgaaggat ggcacgcccg cccgcgacca
     1681 catgcacgtg ccgttcacgc ccatcctaga tggacggttc aactacaagc agatgtgccc
     1741 ccgcagcttc taccggagca tgcacaagga gcttggggat tacctagagg ggcgcatggg
     1801 ctaccgccct gcaatcgagc ttgacgagga gacgcgggcg cagcgtgtat acacggacaa
     1861 gtccgtggat atcgacaagg tacgctgtgc ggtcgataag gccgtggttg aacctgcgaa
     1921 ggacgaagcg gcgcgaatcg tcgcggcggc tagggaagaa gccgccgctt tactgaatga
     1981 tgccgaaacg cgtaaggcag agcttgttac cgagattgcc gacaaggagg gtgaccttgc
     2041 cgagcttgac agccagctcg aggacgtgaa gctcgatatc gaagatgagc aagaccgatt
     2101 ggagtgtctt cggcaacgag cggacggagt tgcgcgagac attggagagc ttcaacccat
     2161 cgctaccgag gttcggggct gggaagctgc gggaaaagct gagcgcggcg caatcctcga
     2221 taacatcgtt gttaagtgcg atggactcgc gagccgcatc agagcaggag ttgcgggaat
     2281 gcgaatcaaa gtggaagagc ttagaagtcg aatatcgcgc ccgctcgaat atctgatgcg
     2341 cagggaacag ccaaggcaac agagccttaa tgacgtgctg cgcgatgcgc agcgggcggc
     2401 ggatgcgtgg aaccgcgacc acgctgcacc gcagaggacg tatcgcgggc gggctagatg
     2461 atgtgttata gtagtacctg taatacacga caaggaggtg cgacgcgaaa aaaccccgct
     2521 gaagcggggt tttttgcgga gcttcgcttc cacgatgcgc atgattcgga ggccccggcg
     2581 atcgccgggg cctccgtgcg ttgtgtgtcg atttgggatg cagccagatg ggcttcctcg
     2641 gcgcgaaccg gccgagattt ccgaatgcag acggtctcgc ttggacgatg cgctttcgaa
     2701 taatcccggc ggagcgggca ttccagtctt cttatcaata tatctataga atgggcaaag
     2761 cataaaaact tgcatggact aatgcttgaa acccaggaca ataaccttat agcttgtaaa
     2821 ttctatcata attgtggttt caaaatcggc tccgtcgata ctatgttata cgccaacttt
     2881 gaaaacaact ttgaaaaagc tgttttctgg tatttaaggt tttagaatgc aaggaacagt
     2941 gaattggagt tcgtcttgtt ataattagct tcttggggta tctttaaata ctgtagaaaa
     3001 gaggaaggaa ataataaatg gctaaaatga gaatatcacc ggaattgaaa aaactgatcg
     3061 aaaaataccg ctgcgtaaaa gatacggaag gaatgtctcc tgctaaggta tataagctgg
     3121 tgggagaaaa tgaaaaccta tatttaaaaa tgacggacag ccggtataaa gggaccacct
     3181 atgatgtgga acgggaaaag gacatgatgc tatggctgga aggaaagctg cctgttccaa
     3241 aggtcctgca ctttgaacgg catgatggct ggagcaatct gctcatgagt gaggccgatg
     3301 gcgtcctttg ctcggaagag tatgaagatg aacaaagccc tgaaaagatt atcgagctgt
     3361 atgcggagtg catcaggctc tttcactcca tcgacatatc ggattgtccc tatacgaata
     3421 gcttagacag ccgcttagcc gaattggatt acttactgaa taacgatctg gccgatgtgg
     3481 attgcgaaaa ctgggaagaa gacactccat ttaaagatcc gcgcgagctg tatgattttt
     3541 taaagacgga aaagcccgaa gaggaacttg tcttttccca cggcgacctg ggagacagca
     3601 acatctttgt gaaagatggc aaagtaagtg gctttattga tcttgggaga agcggcaggg
     3661 cggacaagtg gtatgacatt gccttctgcg tccggtcgat cagggaggat atcggggaag
     3721 aacagtatgt cgagctattt tttgacttac tggggatcaa gcctgattgg gagaaaataa
     3781 aatattatat tttactggat gaattgtttt agatcgttaa gggatcaact ttgggagaga
     3841 gttcaaaatt gatccttttt ttataacagg aattaaataa taaaaaagcc ggattaataa
     3901 tctggctttt tatattctct ctctagtata taaacgcaga aaggcccacc cgaaggtgag
     3961 ccagtgtgag tcgactagta tttcaacccg cactccccat gcggggagtg actccagaga
     4021 aacaggatga cgccctctcc aagcccattt caacccgcac tccccatgcg gggagtgaca
     4081 ttagttaatc ataatacaaa cagaccagat tgtctgtttg ttggccgcaa cactcgtaca
     4141 aacagaccag attgtctgtt tgttactaca tttattgtac aacacgagcc catttttgtc
     4201 aaataaaatt taaattatat caacgttaat aagacgttgt caataaaatt attttgacaa
     4261 aatagaatag taatacagga tccgaatcgt ttcagttgcc taaataataa aaaagccgga
     4321 ttaataatca aataaccgtt ccttcaatgt tttttttaga aaatttagtt ataatattag
     4381 atatgatact tttaaatatc taattcaagc ttcaaaaaac accaacttag ttcggtggat
     4441 aaacaaagga gtggttatta ttcaaattgc agatcaagct ttagtaaaaa aaatggtgat
     4501 catgagtcca aagagaagaa cacaggcaga gcgcgcaatg gagacccagg gcaagttgat
     4561 tgcagcggcc ctgggggttt tacgggaaaa aggttacgcg ggattccgga tcgcagatgt
     4621 gcccggtgct gctggtgtct ccagaggagc gcagagccat catttcccga caaagcttga
     4681 gcttctgctt gccacttttg aatggcttta cgaacagatc accgaacgca gtcgggctcg
     4741 attagcgaaa ttgaagccag aggatgacgt catccagcaa atgctggacg acgccgccga
     4801 atttttcctc gacgatgact tctctatcag ccttgatttg attgtggctg ccgaccggga
     4861 cccagcgtta cgcgagggta ttcagcgcac ggtagagagg aatcggtttg tcgtcgagga
     4921 tatgtggctt ggtgttctgg tgagccgtgg tctttcgcgt gatgatgcag aagatatcct
     4981 ttggttgata ttcaattcgg tgcgtgggct tgctgttcgt agcctatggc agaaggacaa
     5041 agaacgcttt gagcgtgtca ggaactcgac actcgaaatt gcgcgagagc ggtacgcgaa
     5101 attcaagcgc tagatctaga agcggccgcg aattcagaat agtaatacag gatccgaatc
     5161 gtttcagttg ccgccggatt aataatcaaa taaccgttcc taaaaaatgc cctcttgggt
     5221 tatcaagagg gtcattatat ttcgcaaaaa accccgcttc ggcggggttt tttcgccttg
     5281 aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     5341 ttcttagacg tcaggtggca ctttcgggga aatgtggctt gtccgccttg accgcagcat
     5401 cgaccgtcgt gcgacacgag tcatcgtagt tggacagctc cgaagcgccg ccccattgca
     5461 cgaacacgct cggccggtcg accgtggcca tccagctgta cgcacgagcc gacaccatac
     5521 cggtggccac atggcgggga cccttgcaaa tctggtacgc ctgatgggtg ttcgccgtgt
     5581 tcagaatgcc ctgcatgccg agcgcgctcg ccatgcagta gatgcggctg gtgccgcaca
     5641 tcgagaacat ggagttcccg ccgcgctgct ccttgatctc gttgagcttc gtgccggctt
     5701 cggcgagagc ctcgtcccag ctgatgcgca cccaacccgg atcgtcgtcg cccttgggat
     5761 tcgtgcgttt catgggatac gcgagccgat cgggatggta gcacgcctgc agcgaagcct
     5821 gcgacttgct gcagctgttc cccgacgacg cgaacgagct ctcgtccccc tcgatcttga
     5881 tggcgcggcc gttctccacc gtcacccaca cgccgcattc catcttgccg cacccgcggc
     5941 acatcgtgcg aatacgctgc acgccgtccg acgacgcaac cgttccgctg tcgtgctctt
     6001 cggcaagcgc cgccggcgcc gttgccgacg tcagcgcaag cgcagcgctc gtcacccctg
     6061 cgagcctcat gaagctgcga cggctcatgg tgccgaacga ccgcctttgc gtggtgttcg
     6121 ccatagctcc tcctctgccg gttccctttt cacccgcact ccaacgagcg ggccggacca
     6181 agcgtccgtg cgaaccatga tgcagatcga gcgcgagttg agcaccccac cgctgatgcg
     6241 aactctgttc aaccaggtca gaaaccctac tatcgagctc atgaaagacc taccaccgag
     6301 agtggtattt cgccatccgc ccgccgccgc ggtatcggcg aattgctata ctgacgacaa
     6361 tgctcgaacg agccttatcg ccatggggat gtgagcatgc tgcacagcca ttacccttgc
     6421 aaccgaagct ctttcagtct ccggaaaaac gtcgagacgc tcacgccgag ccgagccgcc
     6481 gcctcttcgg catccagctc gcgcgcttca tagctcgaaa ccacctcacg cagcacaagt
     6541 tcgtccatcg gcttgcgctt gcggcccttg tacgcgcccc gctcggcagc cgcgcgcacg
     6601 cccgagatgc gcttctcctc gcgcgcgcga tacgccttct cgtacatgga tagaaggacg
     6661 tcgtgcagca ccttgcgctc gctgtcggca ataggctcgc cgttcagaaa gagggcatcg
     6721 cctcggcaca gcgccgaaag caccgctctt gcagcctgct cggagttcgc ggcgcccgcg
     6781 atcgcgtcgg ccacgtcgac atcggcaccg acctcgcgtt cttcaaatgg ccgaacgtac
     6841 aagccatctc ctatgtagcg cgtatccgtc ctgatgagcc attcgatctt gtcgaggtac
     6901 tccatgccgc atgcatcgag gagctgccac aagccctcgc ggctctcgga cggcgcccgc
     6961 tcgctcacga acgtcggggt gacgttctcg cgcacgtatt ccgtcttgcg caacgacagg
     7021 tcgtaccccg gcaccccctg gaaaagcggg ggctcgagca aatcgatcac cggccaattc
     7081 ggaacgaacg tatagcgaaa cgactcgtcc tcgcgcacca ggtacgtgat cgcccccacc
     7141 tcgtatacca ggcccccggg accatcgagg ctgatcatcc ccgtagtcgt cgattccctc
     7201 atgcgcccct cctcaccagg tccgaatagg gggcttccag aatgacccga tagcgcgcat
     7261 cgagcatatg gcagtaaaac tccttgcgcc gatcgcttat gttcggaacg gatcgcacga
     7321 gctgatgagc cgcttccaaa tccatacgct ccatgacctc tcgcagggct cgattacact
     7381 cgtcgaattc caagctggac acgactgtca gaccaagttt actcatatat actttagatt
     7441 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     7501 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     7561 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     7621 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     7681 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     7741 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     7801 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     7861 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     7921 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     7981 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     8041 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     8101 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     8161 aaaaacgcca gcaacgcggc ccgagatgcg ccgcgtgcgg ctgctggaga tggcggacgc
     8221 gatggatatg ttctgccaag ggttggtttg cgcattcaca gttctccgca agaattgatt
     8281 ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc cgccggcatg agctcctcta
     8341 gtagagagcg ttcaccgaca aacaacagat aaaacgaaag gcccagtctt tcgactgagc
     8401 ctttcgtttt atttgaaaaa gaaaagcccc agcaagcttg gcggcagcag
//
