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pbr

Rust drunk on perbase pileups and lua expressions.

An example use is in calculating mutation rates. Variant callers have filters such as these that affect the number of mutations called; we also want to scale the denominator in similar way so we can compare rates across samples.

Expression

The following attributes are available on the read object in lua expressions:

mapping_quality
flags # integer. must use bit32 lua module (builtin here) to do operations
tid
start
stop
# where in the current read is the pileup given by qpos with convenience of distance_from_[left/right]_end
qpos
distance_from_5prime
distance_from_3prime
insert_size
qname
bq # base_quality at current site
length # length of the read sequence
sequence

An example expression could be:

-- high mapping-q    and base-quality for this column
read.mapping_quality > 10 and read.bq > 20 \
--  and  not within 10 bases of left end      or     right end
    and read.distance_from_5prime > 10 and read.distance_from_3prime > 10 \
--  and  exclude read if unmapped, not primary, qc_fail, or duplicate.
    and bit32.band(read.flags, bit32.bor(4, 256, 512, 1024)) == 0 \
--  and exclude read if it has more than 5% N's in the sequence
    and string_count(read.sequence, 'N') < 0.05 * read.length

this runs as:

pbr $bam "return $expression" > out.pileup

where the $expression argument is the lua expression.

  • Note that we can use, e.g. print(read.qname, read.flags); return $expression) to help with debugging.
  • Note that the expression must contain 'return'

Usage

pileups filtered with lua expressions

Usage: pbr [OPTIONS] <BAM_PATH> <EXPRESSION>

Arguments:
  <BAM_PATH>    Path to the bamfile
  <EXPRESSION>  Lua expression to evaluate

Options:
  -t, --threads <THREADS>                  Number of threads to use [default: 2]
  -m, --max-depth <MAX_DEPTH>              maximum depth in the pileup [default: 100000]
  -b, --bedfile <BEDFILE>                  optional path to the BED of include regions
  -f, --fasta <FASTA>                      optional path to the reference fasta file
  -e, --exclude <EXCLUDE>                  optional path to BED of exclude regions
      --mate-fix                           adjust depth to not double count overlapping mates
  -p, --pile-expression <PILE_EXPRESSION>  optional expression required for the pileup
  -h, --help                               Print help
  -V, --version                            Print version

PileExpression

Note that the pile-expression is also a lua expression; it is applied to the Pileup (column) rather than to the reads. The available attributes on the pile object are:

depth,a,c,g,t,n,fail,ins,del,ref_skip

An example --pile-expression would look like:

return pile.n / pile.depth < 0.05

To require that fewer than 5% of the reads in the pile are 'N'. Positions that do not pass this expression will not be printed.