LOCUS       sequence-440440-        3269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA expression vector - pBG28 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3269)
  AUTHORS   Roghair Stroud MN, Vang DX, Halverson LJ
  TITLE     Optimized CRISPR Interference System for Investigating Pseudomonas 
            alloputida Genes Involved in Rhizosphere Microbiome Assembly.
  JOURNAL   ACS Synth Biol. 2024 Aug 20. doi: 10.1021/acssynbio.4c00312.
  PUBMED    39163848
REFERENCE   2  (bases 1 to 3269)
  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: "ACS Synth 
            Biol. 2024 Aug 20. doi: 10.1021/acssynbio.4c00312."
COMMENT     Sequence Label: pSpyD
FEATURES             Location/Qualifiers
     source          1..3269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(56..142)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_RNA        236..311
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      313..356
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      449..543
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(664..683)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             675..1490
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     oriT            1657..1765
                     /note="incP origin of transfer"
     CDS             complement(1776..2438)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      2439..3210
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
ORIGIN
        1 gaagcccgga gagcaagccc gtaggggggg cgcgcccagc tgtctagggc ggcggatttg
       61 tcctactcag gagagcgttc accgacaaac aacagataaa acgaaaggcc cagtctttcg
      121 actgagcctt tcgttttatt tgatgccttt aattaactag gttgacatgg atataatgta
      181 tgtacctagg ccgcggccgc gcgaattcga gctcggtctc atcactaggg tctctgtttt
      241 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
      301 cgagtcggtg ctataaaacg aaaggcccag tctttcgact gagcctttcg ttttataact
      361 aggtacccgg ggatcctcta gagtcgacct gcaggcatgc aagcttgcgg ccgcgtcgtg
      421 actgggaaaa ccctggcgac tagtcttgga ctcctgttga tagatccagt aatgacctca
      481 gaactccatc tggatttgtt cagaacgctc ggttgccgcc gggcgttttt tattggtgag
      541 aatccagggg tccccaataa ttacgattta aatttgtgtc tcaaaatctc tgatgttaca
      601 ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta
      661 atacaagggg tgttatgagc catattcagc gtgaaacgag ctgtagccgt ccgcgtctga
      721 acagcaacat ggatgcggat ctgtatggct ataaatgggc gcgtgataac gtgggtcaga
      781 gcggcgcgac catttatcgt ctgtatggca aaccggatgc gccggaactg tttctgaaac
      841 atggcaaagg cagcgtggcg aacgatgtga ccgatgaaat ggtgcgtctg aactggctga
      901 ccgaatttat gccgctgccg accattaaac attttattcg caccccggat gatgcgtggc
      961 tgctgaccac cgcgattccg ggcaaaaccg cgtttcaggt gctggaagaa tatccggata
     1021 gcggcgaaaa cattgtggat gcgctggccg tgtttctgcg tcgtctgcat agcattccgg
     1081 tgtgcaactg cccgtttaac agcgatcgtg tgtttcgtct ggcccaggcg cagagccgta
     1141 tgaacaacgg cctggtggat gcgagcgatt ttgatgatga acgtaacggc tggccggtgg
     1201 aacaggtgtg gaaagaaatg cataaactgc tgccgtttag cccggatagc gtggtgaccc
     1261 acggcgattt tagcctggat aacctgattt tcgatgaagg caaactgatt ggctgcattg
     1321 atgtgggccg tgtgggcatt gcggatcgtt atcaggatct ggccattctg tggaactgcc
     1381 tgggcgaatt tagcccgagc ctgcaaaaac gtctgtttca gaaatatggc attgataatc
     1441 cggatatgaa caaactgcaa tttcatctga tgctggatga atttttctaa taattaattg
     1501 gaccgcggtc cgcgcgttgt ccttttccgc tgcataaccc tgcttcgggg tcattatagc
     1561 gattttttcg gtatatccat cctttttcgc acgatataca ggattttgcc aaagggttcg
     1621 tgtagacttt ccttggtgta tccaacggcg tcagccgggc aggataggtg aagtaggccc
     1681 acccgcgagc gggtgttcct tcttcactgt cccttattcg cacctggcgg tgctcaacgg
     1741 gaatcctgct ctgcgaggct ggccgtaggc cggccctacc ggcgcggcag cgttacccgt
     1801 gtcggcggct ccaacggctc gccatcgtcc agaaaacacg gctcatcggg catcggcagg
     1861 cgctgctgcc cgcgccgttc ccattcctcc gtttcggtca aggctggcag gtctggttcc
     1921 atgcccggaa tgccgggctg gctgggcggc tcctcgccgg ggccggtcgg tagttgctgc
     1981 tcgcccggat acagggtcgg gatgcggcgc aggtcgccat gccccaacag cgattcgtcc
     2041 tggtcgtcgt gatcaaccac cacggcggca ctgaacaccg acaggcgcaa ctggtcgcgg
     2101 ggctggcccc acgccacgcg gtcattgacc acgtaggccg acacggtgcc ggggccgttg
     2161 agcttcacga cggagatcca gcgctcggcc accaagtcct tgactgcgta ttggaccgtc
     2221 cgcaaagaac gtccgatgag cttggaaagt gtcttctggc tgaccaccac ggcgttctgg
     2281 tggcccatct gcgccacgag gtgatgcagc agcattgccg ccgtgggttt cctcgcaata
     2341 agcccggccc acgcctcatg cgctttgcgt tccgtttgca cccagtgacc gggcttgttc
     2401 ttggcttgaa tgccgatttc tctggactgc gtggccatgc ttatctccat gcggtagggg
     2461 tgccgcacgg ttgcggcacc atgcgcaatc agctgcaact tttcggcagc gcgacaacaa
     2521 ttatgcgttg cgtaaaagtg gcagtcaatt acagattttc tttaacctac gcaatgagct
     2581 attgcggggg gtgccgcaat gagctgttgc gtacccccct tttttaagtt gttgattttt
     2641 aagtctttcg catttcgccc tatatctagt tctttggtgc ccaaagaagg gcacccctgc
     2701 ggggttcccc cacgccttcg gcgcggctcc ccctccggca aaaagtggcc cctccggggc
     2761 ttgttgatcg actgcgcggc cttcggcctt gcccaaggtg gcgctgcccc cttggaaccc
     2821 ccgcactcgc cgccgtgagg ctcggggggc aggcgggcgg gcttcgccct tcgactgccc
     2881 ccactcgcat aggcttgggt cgttccaggc gcgtcaaggc caagccgctg cgcggtcgct
     2941 gcgcgagcct tgacccgcct tccacttggt gtccaaccgg caagcgaagc gcgcaggccg
     3001 caggccggag gcttttcccc agagaaaatt aaaaaaattg atggggcaag gccgcaggcc
     3061 gcgcagttgg agccggtggg tatgtggtcg aaggctgggt agccggtggg caatccctgt
     3121 ggtcaagctc gtgggcaggc gcagcctgtc catcagcttg tccagcaggg ttgtccacgg
     3181 gccgagcgaa gcgagccagc cggtggccgc tcgcggccat cgtccacata tccacgggct
     3241 ggcaagggag cgcagcgacc gcgcagggc
//
