Package 'FastqCleaner'

Title: A Shiny Application for Quality Control, Filtering and Trimming of FASTQ Files
Description: An interactive web application for quality control, filtering and trimming of FASTQ files. This user-friendly tool combines a pipeline for data processing based on Biostrings and ShortRead infrastructure, with a cutting-edge visual environment. Single-Read and Paired-End files can be locally processed. Diagnostic interactive plots (CG content, per-base sequence quality, etc.) are provided for both the input and output files.
Authors: Leandro Roser [aut, cre], Fernán Agüero [aut], Daniel Sánchez [aut]
Maintainer: Leandro Roser <[email protected]>
License: MIT + file LICENSE
Version: 1.25.0
Built: 2024-11-29 07:12:10 UTC
Source: https://github.com/bioc/FastqCleaner

Help Index


Remove full and partial adapters from a ShortReadQ object

Description

This program can remove adapters and partial adapters from 3' and 5', using the functions trimLRPatterns The program extends the methodology of the trimLRPatterns function of Biostrings, being also capable of removing adapters present within reads and with other additional otpions (e.g., threshold of minimum number of bases for trimming). For a given position in the read, the two Biostrings functions return TRUE when a match is present between a substring of the read and the adapter. As trimLRPatterns , adapter_filter also selects region and goes up to the end of the sequence in the corresponding flank as the best match. The default error rate is 0.2. If several valid matches are found, the function removes the largest subsequence. Adapters can be anchored or not. When indels are allowed, the second method uses the 'edit distance' between the subsequences and the adapter

Usage

adapter_filter(
  input,
  Lpattern = "",
  Rpattern = "",
  rc.L = FALSE,
  rc.R = FALSE,
  first = c("R", "L"),
  with_indels = FALSE,
  error_rate = 0.2,
  anchored = TRUE,
  fixed = "subject",
  remove_zero = TRUE,
  checks = TRUE,
  min_match_flank = 3L,
  ...
)

Arguments

input

ShortReadQ object

Lpattern

5' pattern (character or DNAString object)

Rpattern

3' pattern (character or DNAString object)

rc.L

Reverse complement Lpattern? default FALSE

rc.R

Reverse complement Rpatter? default FALSE

first

trim first right('R') or left ('L') side of sequences when both Lpattern and Rpattern are passed

with_indels

Allow indels? This feature is available only when the error_rate is not null

error_rate

Error rate (value in the range [0, 1] The error rate is the proportion of mismatches allowed between the adapter and the aligned portion of the subject. For a given adapter A, the number of allowed mismatches between each subsequence s of A and the subject is computed as: error_rate * L_s, where L_s is the length of the subsequence s

anchored

Adapter or partial adapter within sequence (anchored = FALSE, default) or only in 3' and 5' terminals? (anchored = TRUE)

fixed

Parameter passed to trimLRPatterns Default 'subject', ambiguities in the pattern only are interpreted as wildcard. See the argument fixed in trimLRPatterns

remove_zero

Remove zero-length sequences? Default TRUE

checks

Perform checks? Default TRUE

min_match_flank

Do not trim in flanks of the subject, if a match has min_match_flank of less length. Default 1L (only trim with >=2 coincidences in a flank match)

...

additional parameters passed to trimLRPatterns

Value

Edited DNAString or DNAStringSet object

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create 6 sequences of width 43
set.seed(10)
input <- random_seq(6, 43)

# add adapter in 3' 
adapter <- "ATCGACT"

input <- paste0(input, as.character(DNAString(adapter)))
input <- DNAStringSet(input)

# create qualities of width 50
set.seed(10)
input_q <- random_qual(c(30,40), slength = 6, swidth = 50, 
encod = 'Sanger')

# create names
input_names <- seq_names(length(input))

# create ShortReadQ object
my_read <- ShortReadQ(sread = input, quality = input_q, id = input_names)

# trim adapter
filtered <- adapter_filter(my_read, Rpattern = adapter)

# look at the filtered sequences
sread(filtered)

Check quality encoding

Description

Check quality encoding

Usage

check_encoding(x = NULL, custom = NULL)

Arguments

x

Quality values

custom

custom encoding from the following:

'Sanger' ——–> expected range: [0, 40]

'Illumina1.8' ——–> expected range: [0, 41]

'Illumina1.5' ——–> expected range: [0, 40]

'Illumina1.3' ——–> expected range: [3, 40]

'Solexa' ——–> expected range: [-5, 40]

Value

List with encoding information

Author(s)

Leandro Roser [email protected]

Examples

require(Biostrings)

x <- list(PhredQuality(0:40), SolexaQuality(-5:40), IlluminaQuality(3:40))
x <- lapply(x, function(i)utf8ToInt(as.character(i)[1]))
lapply(x, check_encoding)
 
SolexaQuality(0:40)
IlluminaQuality(0:40)

Remove sequences with low complexity

Description

The program removes low complexity sequences, computing the entropy with the observed frequency of dinucleotides.

Usage

complex_filter(input, threshold = 0.5, referenceEntropy = 3.908135)

Arguments

input

ShortReadQ object

threshold

A threshold value computed as the relation of the H of the sequences and the reference H. Default is 0.5

referenceEntropy

Reference entropy. By default, the program uses a value of 3.908, that corresponds to the entropy of the human genome in bits

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create  sequences of different width
set.seed(10)
input <- lapply(c(0, 6, 10, 16, 20, 26, 30, 36, 40), 
               function(x) random_seq(1, x))


# create repetitive 'CG' sequences with length adequante 
# for a total length:
# input +  CG = 40

set.seed(10)
CG <- lapply(c(20, 17, 15, 12, 10, 7, 5, 2, 0), 
            function(x) paste(rep('CG', x), collapse = ''))


# concatenate input and CG
input  <- mapply('paste', input, CG, sep = '')
input <- DNAStringSet(input)

# plot relative entropy (E, Shannon 1948)

freq <- dinucleotideFrequency(input)
freq  <- freq /rowSums(freq)
H <- -rowSums(freq  * log2(freq), na.rm = TRUE)
H_max <- 3.908135  # max entropy
plot(H/H_max, type='b', xlab = 'Sequence', ylab= 'E')


# create qualities of width 40

set.seed(10)
input_q <- random_qual(c(30,40), slength = 9, swidth = 40, 
                       encod = 'Sanger')

# create names
input_names <- seq_names(9)

# create ShortReadQ object
my_read <- ShortReadQ(sread = input, quality = input_q, id = input_names)

# apply the filter
filtered <- complex_filter(my_read)

# look at the filtered sequences
sread(filtered)

Remove a fixed number of bases of a ShortReadQ object from 3' or 5'

Description

The program removes a given number of bases from the 3' or 5' regions of the sequences contained in a ShortReadQ object

Usage

fixed_filter(input, trim3 = NA, trim5 = NA)

Arguments

input

ShortReadQ object

trim3

Number of bases to remove from 3'

trim5

Number of bases to remove from 5'

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create 6 sequences of width 20
 
set.seed(10)
input <- random_seq(6, 20)

# create qualities of width 20 

set.seed(10)
input_q <- random_qual(c(30,40), slength = 6, swidth = 20, 
encod = 'Sanger')


# create names
input_names <- seq_names(6)

# create ShortReadQ object
my_read <- ShortReadQ(sread = input, quality = input_q, id = input_names)

# apply the filter 
filtered3 <- fixed_filter(my_read, trim5 = 5)

filtered5 <- fixed_filter(my_read, trim3 = 5)

filtered3and5 <- fixed_filter(my_read, trim3 = 10, trim5 = 5)

# look at the trimmed sequences
sread(filtered3)
sread(filtered5)
sread(filtered3and5)

Inject a letter in a set of sequences at random positions

Description

Inject a letter in a set of sequences at random positions

Usage

inject_letter_random(
  my_seq,
  how_many_seqs = NULL,
  how_many_letters = NULL,
  letter = "N"
)

Arguments

my_seq

character vector with sequences to inject

how_many_seqs

How many sequences pick to inject Ns. An interval [min_s, max_s] with min_s minimum and max_s maximum sequences can be passed. In this case, a value is picked from the interval. If NULL, a random value within the interval [1, length(my_seq)] is picked.

how_many_letters

How many times inject the letter in the i sequences that are going to be injected. An interval [min_i max_i] can be passed. In this case, a value is randomly picked for each sequence i. This value represents the number of times that the letter will be injected in the sequence i. If NULL, a random value within the interval [1, width(my_seq[i])] is picked for each sequence i.

letter

Letter to inject. Default: 'N'

Value

character vector

Author(s)

Leandro Roser [email protected]

Examples

# For reproducible examples, make a call to set.seed before 
# running each random function

set.seed(10)
s <- random_seq(slength = 10, swidth = 20)

set.seed(10)
s <- inject_letter_random(s, how_many_seqs = 1:30, how_many= 2:10)

Launch FastqCleaner application

Description

Launch FastqCleaner application

Usage

launch_fqc(launch.browser = TRUE, ...)

Arguments

launch.browser

Launch in browser? Default TRUE

...

Additional parameters passed to runApp

Value

Launch the application, without return value

Author(s)

Leandro Roser [email protected]

Examples

# Uncomment and paste in te console to launch the application:
# launch_fqc() 

NULL

Filter sequences of a FASTQ file by length

Description

The program removes from a ShortReadQ object those sequences with a length lower than rm.min or/and higher than rm.max

Usage

length_filter(input, rm.min = NA, rm.max = NA)

Arguments

input

ShortReadQ object

rm.min

Threshold value for the minimun number of bases

rm.max

Threshold value for the maximum number of bases

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create  ShortReadQ object width widths between 1 and 100
 
 set.seed(10)
input <- random_length(100, widths = 1:100)

# apply the filter, removing sequences length < 10 or length > 80
filtered <- length_filter(input, rm.min = 10, rm.max = 80)

# look at the filtered sequences
sread(filtered)

Remove sequences with non-identified bases (Ns) from a ShortReadQ object

Description

This program is a wrapper to nFilter. It removes the sequences with a number of N's above a threshold value 'rm.N'. All the sequences with a number of N > rm.N (N >= rm.N) will be removed

Usage

n_filter(input, rm.N)

Arguments

input

ShortReadQ object

rm.N

Threshold value of N's to remove a sequence from the output (sequences with number of Ns > threshold are removed) For example, if rm.N is 3, all the sequences with a number of Ns > 3 (Ns >= 4) will be removed

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create 6 sequences of width 20
set.seed(10)
input <- random_seq(50, 20)

# inject N's
set.seed(10)
input <- inject_letter_random(input, how_many_seqs = 1:30, 
how_many = 1:10)

input <- DNAStringSet(input)


# watch the N's frequency
hist(letterFrequency(input, 'N'), breaks = 0:10, 
main  = 'Ns Frequency', xlab = '# Ns')

# create qualities of width 20
set.seed(10) 
input_q <- random_qual(50, 20)

# create names
input_names <- seq_names(50)

# create ShortReadQ object
my_read <- ShortReadQ(sread = input, quality = input_q, id = input_names)

# apply the filter 
filtered <- n_filter(my_read, rm.N = 3)

# watch the filtered sequences
sread(filtered)

# watch the N's frequency
hist(letterFrequency(sread(filtered), 'N'), 
main = 'Ns distribution', xlab = '')

Filter sequences by their average quality

Description

The program removes the sequences with a quality lower the 'minq' threshold

Usage

qmean_filter(input, minq, q_format = NULL, check.encod = TRUE)

Arguments

input

ShortReadQ object

minq

Quality threshold

q_format

Quality format used for the file, as returned by check.encoding

check.encod

Check the encoding of the sequence? This argument is incompatible with q_format

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require(ShortRead)

set.seed(10)
# create 30 sequences of width 20
input <- random_seq(30, 20)

# create qualities of width 20 
## high quality (15 sequences)
set.seed(10)
my_qual <- random_qual(c(30,40), slength = 15, swidth = 20,
                       encod = 'Sanger')
## low quality (15 sequences)
set.seed(10)
my_qual_2 <-   random_qual(c(5,30), slength = 15, swidth = 20,
                           encod = 'Sanger')

# concatenate vectors
input_q<- c(my_qual, my_qual_2)

# create names
input_names <- seq_names(30)

# create ShortReadQ object
my_read <- ShortReadQ(sread = input, quality = input_q, id = input_names)

# watch the average qualities
alphabetScore(my_read) / width(my_read)

# apply the filter
filtered <- qmean_filter(my_read, minq = 30)

# watch the average qualities
alphabetScore(my_read) / width(my_read)


# watch the filtered sequences
sread(filtered)

Create a named object with random sequences and qualities

Description

Create a ShortReadQ object with random sequences and qualities

Usage

random_length(
  n,
  widths,
  random_widths = TRUE,
  replace = TRUE,
  len_prob = NULL,
  seq_prob = c(0.25, 0.25, 0.25, 0.25),
  q_prob = NULL,
  nuc = c("DNA", "RNA"),
  qual = NULL,
  encod = c("Sanger", "Illumina1.8", "Illumina1.5", "Illumina1.3", "Solexa"),
  base_name = "s",
  sep = "_"
)

Arguments

n

number of sequences

widths

width of the sequences

random_widths

width must be picked at random from the passed parameter 'widths', considering the value as an interval where any integer can be picked. Default TRUE. Otherwise, widths are picked only from the vector passed.

replace

sample widths with replacement? Default TRUE.

len_prob

vector with probabilities for each width value. Default NULL (equiprobability)

seq_prob

a vector of four probabilities values to set the frequency of the nucleotides 'A', 'C', 'G', 'T', for DNA, or 'A', 'C', 'G', 'U', for RNA. For example = c(0.25, 0.25, 0.5, 0). Default is = c(0.25, 0.25, 0.25, 0.25) (equiprobability for the 4 bases). If the sum of the probabilities is > 1, the values will be nomalized to the range [0, 1].

q_prob

a vector of range = range(qual), with probabilities to set the frequency of each quality value. Default is equiprobability. If the sum of the probabilities is > 1, the values will be nomalized to the range [0, 1].

nuc

create sequences of DNA (nucleotides = c('A', 'C', 'G', 'T')) or RNA (nucleotides = c('A, 'C', 'G', 'U'))?. Default: 'DNA'

qual

quality range for the sequences. It must be a range included in the selected encoding:

'Sanger' = [0, 40]

'Illumina1.8' = [0, 41]

'Illumina1.5' = [0, 40]

'Illumina1.3' = [3, 40]

'Solexa' = [-5, 40]

example: for a range from 20 to 30 in Sanger encoding, pass the argument = c(20, 30)

encod

sequence encoding

base_name

Base name for strings

sep

Character separing base names and the read number. Default: '_'

Value

ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

# For reproducible examples, make a call to set.seed before 
# running each random function

set.seed(10)
s1 <- random_seq(slength = 10, swidth = 20)
s1

set.seed(10)
s2 <- random_seq(slength = 10, swidth = 20, 
prob = c(0.6, 0.1, 0.3, 0))
s2

Create random qualities for a given encoding

Description

Create a BStringSet object with random qualities

Usage

random_qual(
  slength,
  swidth,
  qual = NULL,
  encod = c("Sanger", "Illumina1.8", "Illumina1.5", "Illumina1.3", "Solexa"),
  prob = NULL
)

Arguments

slength

number of sequences

swidth

width of the sequences

qual

quality range for the sequences. It must be a range included in the selected encoding:

'Sanger' = [0, 40]

'Illumina1.8' = [0, 41]

'Illumina1.5' = [0, 40]

'Illumina1.3' = [3, 40]

'Solexa' = [-5, 40]

example: for a range from 20 to 30 in Sanger encoding, pass the argument = c(20, 30)

encod

sequence encoding

prob

a vector of range = range(qual), with probabilities to set the frequency of each quality value. Default is equiprobability. If the sum of the probabilities is > 1, the values will be nomalized to the range [0, 1].

Value

BStringSet object

Author(s)

Leandro Roser [email protected]

Examples

q <- random_qual(30, 20)
q

Create random sequences

Description

Create a DNAStringSet object with random sequences

Usage

random_seq(
  slength,
  swidth,
  nuc = c("DNA", "RNA"),
  prob = c(0.25, 0.25, 0.25, 0.25)
)

Arguments

slength

Number of sequences

swidth

Width of the sequences

nuc

Create sequences of DNA (nucleotides = c('A', 'C', 'G', 'T')) or RNA (nucleotides = c('A, 'C', 'G', 'U'))?. Default: 'DNA'

prob

A vector of four probability values used to set the frequency of the nucleotides 'A', 'C', 'G', 'T', for DNA, or 'A', 'C', 'G', 'U', for RNA. For example = c(0.25, 0.25, 0.5, 0). Default is = c(0.25, 0.25, 0.25, 0.25) (equiprobability for the 4 bases). If the sum of the probabilities is > 1, the values will be nomalized to the range [0, 1].

Value

DNAStringSet object

Author(s)

Leandro Roser [email protected]

Examples

# For reproducible examples, make a call to set.seed before 
# running each random function

set.seed(10)
s1 <- random_seq(slength = 10, swidth = 20)
s1

set.seed(10)
s2 <- random_seq(slength = 10, swidth = 20, 
prob = c(0.6, 0.1, 0.3, 0))
s2

Remove a set of sequences

Description

Removes a set of sequences

Usage

seq_filter(input, rm.seq)

Arguments

input

ShortReadQ object

rm.seq

Ccharacter vector with sequences to remove

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require(ShortRead)

set.seed(10)
input <- random_length(30, 3:7)
rm.seq  = c('TGGTC', 'CGGT', 'GTTCT', 'ATA')

# verify that some sequences match
match_before <- unlist(lapply(rm.seq,
 function(x) grep(x, as.character(sread(input)))))

filtered <- seq_filter(input,rm.seq =  rm.seq)

# verify that matching sequences were removed
match_after <- unlist(lapply(rm.seq, 
function(x) grep(x, as.character(sread(filtered)))))

Create sequences names

Description

Create BStringSet object with names

Usage

seq_names(n, base_name = "s", sep = "_")

Arguments

n

Number of reads

base_name

Base name for strings

sep

Character separing base names and the read number. Default: '_

Value

BStringSet object

Examples

snames <- seq_names(10)
snames
snames2 <- seq_names(10, base_name = 's', sep = '.')
snames2

Filter sequences with low quality in 3' tails

Description

The program removes from the 3' tails of the sequences a set of nucleotides showing a quality < a threshold value in a ShortReadQ object

Usage

trim3q_filter(
  input,
  rm.3qual,
  q_format = NULL,
  check.encod = TRUE,
  remove_zero = TRUE
)

Arguments

input

ShortReadQ object

rm.3qual

Quality threshold for 3' tails

q_format

Quality format used for the file, as returned by check_encoding

check.encod

Check the encoding of the sequence? This argument is incompatible with q_format. Default TRUE

remove_zero

Remove zero-length sequences?

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

# create 6 sequences of width 20
set.seed(10)
input <- random_seq(6, 20)

# create qualities of width 15 and paste to qualities
# of length 5 used for the tails.
# for two of the sequences, put low qualities in tails

set.seed(10)
my_qual <- random_qual(c(30,40), slength = 6, swidth = 15, 
encod = 'Sanger')

set.seed(10)
tails <-   random_qual(c(30,40), slength = 6, swidth = 5, 
 encod = 'Sanger')
 
set.seed(10)
tails[2:3] <- random_qual(c(3, 20), slength = 2, 
swidth = 5,  encod = 'Sanger')
my_qual <- paste0(my_qual, tails)
input_q <- BStringSet(my_qual)
# create names
input_names <- seq_names(6)

# create ShortReadQ object
my_read <- ShortReadQ(sread = input,
quality = input_q, id = input_names)

# apply the filter 
filtered <- trim3q_filter(my_read, rm.3qual = 28)

# look at the trimmed sequences
sread(filtered)

Remove duplicated sequences in a FASTQ file

Description

This program is a wrapper to occurrenceFilter. It removes the duplicated sequences of a FASTQ file.

Usage

unique_filter(input)

Arguments

input

ShortReadQ object

Value

Filtered ShortReadQ object

Author(s)

Leandro Roser [email protected]

Examples

require('Biostrings')
require('ShortRead')

set.seed(10)
s <- random_seq(10, 10)
s <- sample(s, 30, replace = TRUE)
q <- random_qual(30, 10)
n <- seq_names(30)

my_read <- ShortReadQ(sread = s, quality = q, id = n)

# check presence of duplicates
isUnique(as.character(sread(my_read)))

# apply the filter
filtered <- unique_filter(my_read)

isUnique(as.character(sread(filtered)))