find_primespacers {multicrispr} | R Documentation |
Find prime editing spacers around target ranges
find_primespacers( gr, bsgenome, edits = get_plus_seq(bsgenome, gr), nprimer = 13, nrt = 16, ontargetmethod = c("Doench2014", "Doench2016")[1], offtargetmethod = c("bowtie", "pdict")[1], mismatches = 0, nickmatches = 2, indexedgenomesdir = INDEXEDGENOMESDIR, outdir = OUTDIR, verbose = TRUE, plot = TRUE, ... )
gr |
|
bsgenome |
|
edits |
character vector: desired edits on '+' strand.
If named, names should be identical to those of |
nprimer |
n primer nucleotides (default 13, max 17) |
nrt |
n rev transcr nucleotides (default 16, recomm. 10-16) |
ontargetmethod |
'Doench2014' or 'Doench2016': on-target scoring method |
offtargetmethod |
'bowtie' or 'pdict' |
mismatches |
no of primespacer mismatches (default 0, to suppress offtarget analysis: -1) |
nickmatches |
no of nickspacer offtarget mismatches (default 2, to suppresses offtarget analysis: -1) |
indexedgenomesdir |
directory with indexed genomes
(as created by |
outdir |
directory whre offtarget analysis output is written |
verbose |
TRUE (default) or FALSE |
plot |
TRUE (default) or FALSE |
... |
passed to plot_intervals |
Below the architecture of a prime editing site. Edits can be performed anywhere in the revtranscript area.
spacer pam ——————–=== primer revtranscript ————-================ 1..............17....GG.......... .....................CC.......... ———-extension———-
GRanges-class
with prime editing spacer
ranges and following mcols:
* crisprspacer: N20 spacers
* crisprpam: NGG PAMs
* crisprprimer: primer (on PAM strand)
* crisprtranscript: reverse transcript (on PAM strand)
* crisprextension: 3' extension of gRNA
contains: reverse transcription template + primer binding site
sequence can be found on non-PAM strand
* crisprextrange: genomic range of crispr extension
* Doench2016|4: on-target efficiency scores
* off0, off1, off2: number of offtargets with 0, 1, 2 mismatches
* off: total number of offtargets: off = off0 + off1 + ...
* nickrange: nickspacer range
* nickspacer: nickspacer sequence
* nickDoench2016|4: nickspacer Doench scores
* nickoff: nickspacer offtarget counts
find_spacers
to find standard crispr sites
# Find PE spacers for 4 clinically relevant loci (Anzalone et al, 2019) bsgenome <- BSgenome.Hsapiens.UCSC.hg38::BSgenome.Hsapiens.UCSC.hg38 gr <- char_to_granges(c( PRNP = 'chr20:4699600:+', # snp: prion disease HBB = 'chr11:5227002:-', # snp: sickle cell anemia HEXA = 'chr15:72346580-72346583:-', # del: tay sachs disease CFTR = 'chr7:117559593-117559595:+'), # ins: cystic fibrosis bsgenome) spacers <- find_primespacers(gr, bsgenome) spacers <- find_spacers(extend_for_pe(gr), bsgenome, complement = FALSE) # Edit PRNP locus for resistance against prion disease (Anzalone et al, 2019) bsgenome <- BSgenome.Hsapiens.UCSC.hg38::BSgenome.Hsapiens.UCSC.hg38 gr <- char_to_granges(c(PRNP = 'chr20:4699600:+'), bsgenome) find_primespacers(gr, bsgenome) find_primespacers(gr, bsgenome, edits = 'T')