LOCUS       sequence-89764-e         877 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Plasmid encoding multiplexed 4x gRNAs. This is a modified form of 
            the original plasmid described in the paper (Construct 19). mKate2 
            was replaced with dsRed2 because of distribution issues..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 877)
  AUTHORS   Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK
  TITLE     Multiplexed and Programmable Regulation of Gene Networks with an 
            Integrated RNA and CRISPR/Cas Toolkit in Human Cells.
  JOURNAL   Mol Cell. 2014 May 14. pii: S1097-2765(14)00355-4. doi: 
            10.1016/j.molcel.2014.04.022.
  PUBMED    24837679
REFERENCE   2  (bases 1 to 877)
  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: "Mol Cell. 
            2014 May 14. pii: S1097-2765(14)00355-4. doi: 
            10.1016/j.molcel.2014.04.022."
COMMENT     Sequence Label: 
            CMVp-dsRed2-Triplex-28-gRNA3-28-gRNA4-28-gRNA5-28-gRNA6-28-pA
FEATURES             Location/Qualifiers
     source          1..877
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     polyA_signal    43..90
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     115..134
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     protein_bind    complement(339..358)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        complement(367..442)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(475..494)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        complement(503..578)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(611..630)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        complement(639..714)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(747..766)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     misc_RNA        complement(775..850)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 tatcgacaga gtgccagccc tgggaccgaa ccccgcgttt atgaacaaac gacccaacac
       61 cgtgcgtttt attctgtctt tttattgccg tcatagcgcg ggttccttcc ggtattgtct
      121 ccttccgtgt ttcagttagc ctccccctag ggtgggcgaa gaactccagc atgagatccc
      181 cgcgctggag gatcatccag ccggcgtccc ggaaaacgat tccgaagccc aacctttcat
      241 agaaggcggc ggtggaatcg aaatctcgtg atggcaggtt gggcgtcgct tggtcggtca
      301 tttcgaaccc cagagtcccg cttacccggg tttcttagct gcctatacgg cagtgaacga
      361 aaaaaagcac cgactcggtg ccactttttc aagttgataa cggactagcc ttattttaac
      421 ttgctatttc tagctctaaa acgtttccag gagggtgact ccgacttttc ttagctgcct
      481 atacggcagt gaacgaaaaa aagcaccgac tcggtgccac tttttcaagt tgataacgga
      541 ctagccttat tttaacttgc tatttctagc tctaaaacgc tgatggctga cttgatgccc
      601 tgtttcttag ctgcctatac ggcagtgaac gaaaaaaagc accgactcgg tgccactttt
      661 tcaagttgat aacggactag ccttatttta acttgctatt tctagctcta aaacaactgg
      721 ttcttatctg cgtcaccttt tcttagctgc ctatacggca gtgaacgaaa aaaagcaccg
      781 actcggtgcc actttttcaa gttgataacg gactagcctt attttaactt gctatttcta
      841 gctctaaaac gagcacctca gagagtacac gctagct
//
