LOCUS       sequence-206301-        3639 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Guide RNA plasmid targeting lvaA on BBR1-UP origin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3639)
  AUTHORS   Cook TB, Rand JM, Nurani W, Courtney DK, Liu SA, Pfleger BF
  TITLE     Genetic tools for reliable gene expression and recombineering in 
            Pseudomonas putida.
  JOURNAL   J Ind Microbiol Biotechnol. 2018 Jan 3. pii: 
            10.1007/s10295-017-2001-5. doi: 10.1007/s10295-017-2001-5.
  PUBMED    29299733
REFERENCE   2  (bases 1 to 3639)
  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: "J Ind 
            Microbiol Biotechnol. 2018 Jan 3. pii: 10.1007/s10295-017-2001-5. 
            doi: 10.1007/s10295-017-2001-5."
COMMENT     Sequence Label: pgRNAtet-lvaA
FEATURES             Location/Qualifiers
     source          1..3639
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        16..50
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     misc_RNA        71..146
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             167..196
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             212..229
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(285..302)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      455..501
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(554..571)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(706..1368)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVLAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     terminator      complement(1955..2049)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2222..3412)
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTITLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     3041..3060
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        complement(join(3460..3465,3466..3482,3483..3488))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     3572..3590
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3594..3637)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 gaattctaaa gatctttgac agctagctca gtcctaggta taatactagt gtcagtgcta
       61 gctcgagcga gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac
      121 ttgaaaaagt ggcaccgagt cggtgctttt tttgaagctt gggcccgaac aaaaactcat
      181 ctcagaagag gatctgaata gcgccgtcga ccatcatcat catcatcatt gagtttaaac
      241 ggtctccagc ttggctgttt tggcggatga gagaagattt tcagcctgat acagattaaa
      301 tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag cgcggtggtc
      361 ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg
      421 tctccccatg cgagagtagg gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa
      481 agactgggcc tttcgtttta tctgttgttt gtcggtgaac tggatcctta ctcgagtcta
      541 gactgtaggc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
      601 tatcagctca ctcaaaggcg gtaatacggc ctttctgcgt ttatacctag gctacagccg
      661 atagtctgga acagcgcact tacgggttgc tgcgcaaccc aagtgctacc ggcgcggcag
      721 cgtgacccgt gtcggcggct ccaacggctc gccatcgtcc agaaaacacg gctcatcggg
      781 catcggcagg cgctgctgcc cgcgccgttc ccattcctcc gtttcggtca aggctggcag
      841 gtctggttcc atgcccggaa tgccgggctg gctgggcggc tcctcgccgg ggccggtcgg
      901 tagttgctgc tcgcccggat acagggtcgg gatgcggcgc aggtcgccat gccccaacag
      961 cgattcgtcc tggtcgtcgt gatcaaccac cacggcggca ctgaacaccg acaggcgcaa
     1021 ctggtcgcgg ggctggcccc acgccacgcg gtcattgacc acgtaggcca acacggtgcc
     1081 ggggccgttg agcttcacga cggagatcca gcgctcggcc accaagtcct tgactgcgta
     1141 ttggaccgtc cgcaaagaac gtccgatgag cttggaaagt gtcttctggc tgaccaccac
     1201 ggcgttctgg tggcccatct gcgccacgag gtgatgcagc agcattgccg ccgtgggttt
     1261 cctcgcaata agcccggccc acgcctcatg cgctttgcgt tccgtttgca cccagtgacc
     1321 gggcttgttc ttggcttgaa tgccgatttc tctggactgc gtggccatgc ttatctccat
     1381 gcggtagggg tgccgcacgg ttgcggcacc atgcgcaatc agctgcaact tttcggcagc
     1441 gcgacaacaa ttatgcgttg cgtaaaagtg gcagtcaatt acagattttc tttaacctac
     1501 gcaatgagct attgcggggg gtgccgcaat gagctgttgc gtacccccct tttttaagtt
     1561 gttgattttt aagtctttcg catttcgccc tatatctagt tctttggtgc ccaaagaagg
     1621 gcacccctgc ggggttcccc cacgccttcg gcgcggctcc ccctccggca aaaagtggcc
     1681 cctccggggc ttgttgatcg actgcgcggc cttcggcctt gcccaaggtg gcgctgcccc
     1741 cttggaaccc ccgcactcgc cgccgtgagg ctcggggggc aggcgggcgg gcttcgccct
     1801 tcgactgccc ccactcgcat aggcttgggt cgttccaggc gcgtcaaggc caagccgctg
     1861 cgcggtcgct gcgcgagcct tgacccgcct tccacttggt gtccaaccgg caagcgaagc
     1921 gcgcaggccg caggccggag gcactagtgc ttggattctc accaataaaa aacgcccggc
     1981 ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca
     2041 acaggagtcc aagcgagctc tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     2101 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaaagt
     2161 gacaggcaca ttatgcatct tggagtggtg aatccgttag cgaggtgccg ccggcttcca
     2221 ttcaggtcga ggtggcccgg ctccatgcac cgcgacgcaa cgcggggagg cagacaaggt
     2281 atagggcggc gcctacaatc catgccaacc cgttccatgt gctcgccgag gcggcataaa
     2341 tcgccgtgac gatcagcggt ccaatgatcg aagttaggct ggtaagagcc gcgagcgatc
     2401 cttgaagctg tccctgatgg tcgtcatcta cctgcctgga cagcatggcc tgcaacgcgg
     2461 gcatcccgat gccgccggaa gcgagaagaa tcataatggg gaaggccatc cagcctcgcg
     2521 tcgcgaacgc cagcaagacg tagcccagcg cgtcggccgc catgccggcg ataatggcct
     2581 gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca
     2641 agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga aagcggtcct
     2701 cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg ataaagaaga
     2761 cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag ctgactgggt
     2821 tgaaggctct caagggcatc ggtcgacgct ctcccttatg cgactcctgc attaggaagc
     2881 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     2941 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     3001 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
     3061 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
     3121 tccacaggac gggtgtggtc gccatgatcg cgtagtcgat agtggctcca agtagcgaag
     3181 cgagcaggac tgggcggcgg ccaaagcggt cggacagtgc tccgagaacg ggtgcgcata
     3241 gaaattgcat caacgcatat agcgctagca gcacgccata gtgactggcg atgctgtcgg
     3301 aatggacgat atcccgcaag aggcccggca gtaccggcat aaccaagcct atgcctacag
     3361 catccagggt gatggtgccg aggatgacga tgagcgcatt gttagatttc atacacggtg
     3421 cctgactgcg ttagcaattt aactgtgata aactaccgca ttaaagctta tcgatgataa
     3481 gctgtcaaac ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     3541 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc agaatttcag
     3601 ataaaaaaaa tccttagctt tcgctaagga tgatttctg
//
