LOCUS       sequence-307072-        3182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA for ebpA gene (NT) under nisin-inducible nisA promoter, 
            barcoded.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3182)
  AUTHORS   Afonina I, Ong J, Chua J, Lu T, Kline KA
  TITLE     Multiplex CRISPRi System Enables the Study of Stage-Specific Biofilm
            Genetic Requirements in Enterococcus faecalis.
  JOURNAL   mBio. 2020 Oct 20;11(5):e01101-20. doi: 10.1128/mBio.01101-20.
  PUBMED    33082254
REFERENCE   2  (bases 1 to 3182)
  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"; doi: "10.1128/mBio"; 
            journalName: "mBio"; date: "2020-10-20- 20"; volume: "11"; issue: 
            "5"; pages: "e01101-20"
COMMENT     Sequence Label: pGCP123-EbpA_g1
FEATURES             Location/Qualifiers
     source          1..3182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(36..52)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(36..52)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             complement(249..260)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(1901..2695)
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     misc_RNA        3018..3093
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(join(3179..3182,1..17))
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(join(3179..3182,1..15))
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tggcatgctc
       61 gattttttat taaaacgtct caaaatcgtt tctgagacgt tttagcgttt atttcgttta
      121 gttatcggca taatcgttaa aacaggcgtt atcgtagcgt aaaagccctt gagcgtagcg
      181 tggctttgca gcgaagatgt tgtctgttag attatgaaag ccgatgactg aatgaaataa
      241 taagcgcagc gtccttctat ttcggttgga ggaggctcaa gggagtttga gggaatgaaa
      301 ttccctcatg ggtttgattt taaaaattgc ttgcaatttt gccgagcggt agcgctggaa
      361 aatttttgaa aaaaatttgg aatttggaaa aaaatggggg gaaaggaagc gaattttgct
      421 tccgtactac gaccccccat taagtgccga gtgccaattt ttgtgccaaa aacgctctat
      481 cccaactggc tcaagggttt gaggggtttt tcaatcgcca acgaatcgcc aacgttttcg
      541 ccaacgtttt ttataaatct atatttaagt agctttattg ttgtttttat gattacaaag
      601 tgatacacta attttataaa attatttgat tggagttttt taaatggtga tttcagaatc
      661 gaaaaaaaga gttatgattt ctctgacaaa agagcaagat aaaaaattaa cagatatggc
      721 gaaacaaaaa ggtttttcaa aatctgcggt tgcggcgtta gctatagaag aatatgcaag
      781 aaaggaatca gaacaaaaaa aataagcgaa agctcgcgtt tttagaagga tacgagtttt
      841 cgctacttgt ttttgataag gtaattatat catggctatt aaaaatacta aagctagaaa
      901 ttttggattt ttattatatc ctgactcaat tcctaatgat tggaaagaaa aattagagag
      961 tttgggcgta tctatggctg tcagtccttt acacgatatg gacgaaaaaa aagataaaga
     1021 tacatggaat agtagtgatg ttatacgaaa tggaaagcac tataaaaaac cacactatca
     1081 cgttatatat attgcacgaa atcctgtaac aatagaaagc gttaggaaca agattaagcg
     1141 aaaattgggg aatagttcag ttgctcatgt tgagatactt gattatatca aaggttcata
     1201 tgaatatttg actcatgaat caaaggacgc tattgctaag aataaacata tatacgacaa
     1261 aaaagatatt ttgaacatta atgattttga tattgaccgc tatataacac ttgatgaaag
     1321 ccaaaaaaga aaattgaaga atttactttt agatatagtg gatgactata atttggtaaa
     1381 tacaaaagat ttaatggctt ttattcgcct taggggagcg gagtttggaa ttttaaatac
     1441 gaatgatgta aaagatattg tttcaacaaa ctctagcgcc tttagattat ggtttgaggg
     1501 caattatcag tgtggatata gagcaagtta tgcaaaggtt cttgatgctg aaacggggga
     1561 aataaaatga caaacaaaga aaaagagtta tttgctgaaa atgaggaatt aaaaaaagaa
     1621 attaaggact taaaagagcg tattgaaaga tacagagaaa tggaagttga attaagtaca
     1681 acaatagatt tattgagagg agggattatt gaataaataa aagcccccct gacgaaagtc
     1741 gacgatacaa attcctcgta ggcgctcggg acccctatct agcgaacttt tagaaaagat
     1801 ataaaacatc agagtatgga cagttgcgga tgtacttcag aaaagattag atgtctaaaa
     1861 agcttgtagt taaagctttt tagacatcta aatctaggta ctaaaacaat tcatccagta
     1921 aaatataata ttttattttc tcccaatcag gcttgatccc cagtaagtca aaaaatagct
     1981 cgacatactg ttcttccccg atatcctccc tgatcgaccg gacgcagaag gcaatgtcat
     2041 accacttgtc cgccctgccg cttctcccaa gatcaataaa gccacttact ttgccatctt
     2101 tcacaaagat gttgctgtct cccaggtcgc cgtgggaaaa gacaagttcc tcttcgggct
     2161 tttccgtctt taaaaaatca tacagctcgc gcggatcttt aaatggagtg tcttcttccc
     2221 agttttcgca atccacatcg gccagatcgt tattcagtaa gtaatccaat tcggctaagc
     2281 ggctgtctaa gctattcgta tagggacaat ccgatatgtc gatggagtga aagagcctga
     2341 tgcactccgc atacagctcg ataatctttt cagggctttg ttcatcttca tactcttccg
     2401 agcaaaggac gccatcggcc tcactcatga gcagattgct ccagccatca tgccgttcaa
     2461 agtgcaggac ctttggaaca ggcagctttc cttccagcca tagcatcatg tccttttccc
     2521 gttccacatc ataggtggtc cctttatacc ggctgtccgt catttttaaa tataggtttt
     2581 cattttctcc caccagctta tataccttag caggagacat tccttccgta tcttttacgc
     2641 agcggtattt ttcgatcagt tttttcaatt ccggtgatat tctcatttta gccatttatt
     2701 atttccttcc tcttttctac agtatttaaa gataccccaa gaagctaatt ataacaagac
     2761 gaactccaat tcactgttcc ttgcattcta aaaccttaaa taccagaaaa cagctttttc
     2821 aaagttgttt tcaaagttgg cgtataacat agtatcgacg gaagatcctg cagagatcta
     2881 tctaaaacag tcttaattct atcttgagaa agtattggta ataatattat tgtcgataac
     2941 gcgagcataa taaacggctc tgattaaatt ctgaagtttg ttagatacaa tgatttcacg
     3001 ccaggtgctt ttcccgagtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
     3061 tatcaacttg aaaaagtggc accgagtcgg tgcttttttt ggatccacta gtggtaccga
     3121 attcacattc cgcaattggc atgcggccgc caccgcggtg gagctccagc ttttgttccc
     3181 tt
//
