Package 'Rqc'

Quality Control Tool for High-Throughput Sequencing Data

Description

Rqc is an optimized tool designed for quality assessment of high-throughput sequencing data. It performs parallel processing of entire files and produces a report, which contains a set of high-resolution images that can be directly used on publications.

Author(s)

Welliton Souza, Benilton Carvalho

Maintainer: Welliton Souza <[email protected]>

Examples

options(device.ask.default = FALSE)
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  rqc(folder, ".fastq.gz", pair=c(1,1), workers=1)

---

Save time storing longer analysis step on disk

Description

This utility function can be used to save time on task that takes long time to complete. A Rda file are written on disk containing only objects setted to keep. If checkpoint function find related Rda file then this Rda will be loaded.

Usage

checkpoint(
  label,
  CODE,
  path = ".",
  overwrite = FALSE,
  verbose = FALSE,
  keep = NULL
)

Arguments

label	name of this code, will create a Rda file with the same name.
CODE	R code.
path	directory to write/load Rda file.
overwrite	Rerun CODE and replace Rda file.
verbose	argument passed to load function
keep	vector of object/variable name to keep. NULL means error.

Value

Nothing.

Note

Experimental function.

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")

---

Detect file format

Description

Detect file format

Usage

detectFileFormat(file)

Arguments

file

file name

Value

FastqFile or BamFiles objects

Examples

folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
files <- list.files(full.names=TRUE, path=folder)
input <- lapply(files, detectFileFormat)
sapply(input, class)

---

Revert codified DNA sequences to original DNA sequences.

Description

This function receives a vector of strings containing codified DNA and returns a vector of string containing original DNA sequences.

Usage

fromRRDNA(rrdnas)

Arguments

rrdnas

Vector of codified DNA (character vector).

Value

Vector of original DNA sequences (character vector).

Note

This function is used internally to restore original DNA sequences stored in RqcResultSet objects (per file top reads).

Author(s)

Welliton Souza

See Also

perFileTopReads

Examples

dna <- "ATCG"
dna.converted <- toRRDNA(dna)
dna.reverted <- fromRRDNA(dna.converted)
all.equal(dna, dna.reverted)

---

Distance matrix of the similarity between the DNA sequences.

Description

This function receives a vector of strings representing codified DNA sequences and returns a integer matrix representing the similarities between all sequences from input vectors.

Usage

matdist(rrdnas)

Arguments

rrdnas

Vector of codified DNA sequences (character vector).

Value

Matrix $n x n$, where $n$ is the length of the largest original DNA sequence.

Note

This function is used internally to compute data for rqcFileHeatmap function.

Author(s)

Welliton Souza

See Also

rqcFileHeatmap

Examples

dna1 <- toRRDNA("atcgn")
dna2 <- toRRDNA("atcga")
matdist(c(dna1, dna2))

---

Main Rqc function

Description

Rqc is an optimized tool designed for quality assessment of high-throughput sequencing data. It performs parallel processing of entire files and produces an HTML report, which contains a set of high-resolution images that can be directly used on publications.

Usage

rqc(
  path = ".",
  pattern,
  sample = TRUE,
  n = 1e+06,
  group = NULL,
  top = 10,
  pair = NULL,
  outdir = tempdir(),
  file = "rqc_report",
  openBrowser = TRUE,
  workers = multicoreWorkers()
)

Arguments

path	directory path that contains input files.
pattern	a regex expression that macthes to input file names
sample	it reads a random sample from files if this parameter is TRUE.
n	number of sequences to read from each input file. This represents sample size if 'sample' parameter is TRUE, if not represents the chunk size to read on each iteration. By default, it reads a sample of one million sequences from each input file.
group	group name for each input file.
top	number of top over-represented reads. Default is 10 reads.
pair	combination of files for paired-end reads. By default, all input files are treated as single-end. For paired-end, please define a vector of numbers where two index with the same value represent a pair. Examples, single-end c(1,2,3,4) and paired-end c(1,1,2,2).
outdir	output directory path. Is created a temporary directory by default.
file	output file name.
openBrowser	if TRUE opens report file on default Internet Browser.
workers	Number of parallel workers. Set 1 to serial. Default value from multicoreWorkers.

Value

A invisible named list of RqcResultSet objects, each one represents a file.

Author(s)

Welliton Souza

See Also

rqcQA

Examples

options(device.ask.default = FALSE)
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  rqc(folder, ".fastq.gz", pair=c(1,1), workers=1, openBrowser=FALSE)

---

Per cycle average quality by files

Description

This function plots line graph of per cycle average quality.

Usage

rqcCycleAverageQualityCalc(rqcResultSet)

rqcCycleAverageQualityPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

ggplot2 object

Functions

• rqcCycleAverageQualityCalc: calculates necessary statistics

Author(s)

Welliton Souza

See Also

rqcGroupCycleAverageQualityPlot plots cycle-specific quality by groups

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcCycleAverageQualityPlot(rqcResultSet)

---

Biplot of PCA of per cycle read average quality

Description

This function creates a Biplot of PCA of per cycle read average quality

Usage

rqcCycleAverageQualityPcaCalc(rqcResultSet)

rqcCycleAverageQualityPcaPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcCycleAverageQualityPcaCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcCycleAverageQualityPcaPlot(rqcResultSet)

---

Per cycle base calls plot

Description

Creates a bar graph of per cycle base calls.

Usage

rqcCycleBaseCallsCalc(rqcResultSet)

rqcCycleBaseCallsLinePlot(rqcResultSet)

rqcCycleBaseCallsPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcCycleBaseCallsCalc: calculates necessary statistics

• rqcCycleBaseCallsLinePlot: creates a line graph

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcCycleBaseCallsPlot(rqcResultSet)

---

Per cycle percentual GC plot

Description

Creates a line graph of per cycle percentual GC.

Usage

rqcCycleGCCalc(rqcResultSet)

rqcCycleGCPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcCycleGCCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcCycleGCPlot(rqcResultSet)

---

Per cycle quality box plot

Description

Plots per cycle quality box plot.

Usage

rqcCycleQualityBoxCalc(rqcResultSet)

rqcCycleQualityBoxPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcCycleQualityBoxCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcCycleQualityBoxPlot(rqcResultSet)

---

Per cycle quality plot

Description

Creates a graph of per cycle quality.

Usage

rqcCycleQualityCalc(rqcResultSet)

rqcCycleQualityPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcCycleQualityCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, workers=1)
}, keep="rqcResultSet")
rqcCycleQualityPlot(rqcResultSet)

---

Heatmap of distance matrix of top over-represented reads

Description

This function plots a heatmap of distance matrix of top over-represented reads. This function does not work with list of RqcResultSet objects, only with one RqcResultSet object.

Usage

rqcFileHeatmap(
  rqcResultSet,
  dist.method = "euclidean",
  hclust.method = "ward.D"
)

Arguments

rqcResultSet	RqcResultSet object created by rqc and rqcQA functions.
dist.method	the distance measure to be used by dist function.
hclust.method	the agglomeration method to be used by hclust function.

Value

Plot object from ggplot function.

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcFileHeatmap(rqcResultSet[[1]])

---

Per group average quality across cycles

Description

This function plots cycle-specific quality by groups

Usage

rqcGroupCycleAverageQualityCalc(rqcResultSet)

rqcGroupCycleAverageQualityPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

ggplot2 object

Functions

• rqcGroupCycleAverageQualityCalc: calculates necessary statistics

Author(s)

Welliton Souza

See Also

rqcCycleAverageQualityPlot plots cycle-specific quality by files

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcGroupCycleAverageQualityPlot(rqcResultSet)

---

Quality Assessment Rqc function

Description

Process a set of files and returns a list of quality control data. Files must be FASTQ format, compressed or not.

Usage

rqcQA(
  x,
  sample = TRUE,
  n = 1e+06,
  group = rep("None", length(x)),
  top = 10,
  pair = seq_along(x),
  ...
)

## S4 method for signature 'list'
rqcQA(x, sample, n, group, top, pair, workers = multicoreWorkers())

## S4 method for signature 'character'
rqcQA(
  x,
  sample = TRUE,
  n = 1e+06,
  group = rep("None", length(x)),
  top = 10,
  pair = seq_along(x),
  workers = multicoreWorkers()
)

## S4 method for signature 'BamFile'
rqcQA(x, sample, n, group, top, pair)

## S4 method for signature 'FastqFile'
rqcQA(x, sample, n, group, top, pair)

Arguments

x	input file(s)
sample	It reads a random sample from files if this parameter is TRUE.
n	Number of sequences to read from each input file. This represents sample size if 'sample' parameter is TRUE, if not represents the chunk size to read on each iteration. Default is read a sample of one million sequences from each input file.
group	group name for each input file.
top	number of top over-represented reads. Default is 10 reads.
pair	combination of files for paired-end reads. By default, all input files are treated as single-end. For paired-end, please define a vector of numbers where two index with the same value represent a pair. Examples, single-end c(1,2,3,4) and paired-end c(1,1,2,2).
...	other parameters
workers	number of parallel workers

Details

Input files are read using FastStreamer and FastSampler classes of ShortRead package. Process multiple files in parallel using bplapply function of BiocParallel package.

Value

A named list of RqcResultSet objects, each one represents a file.

Methods (by class)

• list: process a list of FastqFile and BamFile objects.

• character: automatically detects file format (using detectFileFormat function) of input files then process.

• BamFile: process only one BAM file.

• FastqFile: process only one FASTQ file.

Author(s)

Welliton Souza

See Also

rqc

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcReadQualityPlot(rqcResultSet)

---

Read frequency plot

Description

This function creates a bar graph of read frequency (in percentage).

Usage

rqcReadFrequencyCalc(rqcResultSet)

rqcReadFrequencyPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcReadFrequencyCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcReadFrequencyPlot(rqcResultSet)

---

Per read mean quality box plot

Description

This function creates crate a graphic charts with box plots describing per read mean quality distribution for each input file

Usage

rqcReadQualityBoxCalc(rqcResultSet)

rqcReadQualityBoxPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcReadQualityBoxCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcReadQualityBoxPlot(rqcResultSet)

---

Per read quality plot

Description

Plots the quality of all the files by read.

Usage

rqcReadQualityCalc(rqcResultSet)

rqcReadQualityPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcReadQualityCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcReadQualityPlot(rqcResultSet)

---

Per read width plot

Description

Creates bar graph of per read width from all elements of input list.

Usage

rqcReadWidthCalc(rqcResultSet)

rqcReadWidthPlot(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet objects created by rqc and rqcQA functions.

Value

Plot object from ggplot function.

Functions

• rqcReadWidthCalc: calculates necessary statistics

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
rqcReadWidthPlot(rqcResultSet)

---

Quality Control HTML Report

Description

Generates an HTML report file.

Usage

rqcReport(
  rqcResultSet,
  outdir = tempdir(),
  file = "rqc_report",
  keepMD = FALSE,
  templateFile = system.file("templates", package = "Rqc", "rqc_report.Rmd")
)

Arguments

rqcResultSet	list of RqcResultSet objects created by rqc and rqcQA functions.
outdir	output directory path. It is created a temporary directory by default.
file	output file name.
keepMD	If true Rqc does not delete markdown file. knit function takes RMarkdown template file (within package) and generates a temporary Markdown file. Next markdownToHTML function takes this markdown file and creates final HTML file.
templateFile	Path of Rmarkdown file as Rqc web report template.

Details

Also creates a directory called "figure" in outdir path.

Value

Report file path.

Author(s)

Welliton Souza

See Also

rqc

rqcQA

Examples

options(device.ask.default = FALSE)
checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
reportFile <- rqcReport(rqcResultSet)
browseURL(reportFile)

---

Class RqcResultSet

Description

Class RqcResultSet

Frequency distribution of cycle-specific base call

Frequency distribution of cycle-specific quality

File information

Top over-represented sequencing reads

Read frequency table

Frequency distribution of per read mean quality

Frequency distribution of read width

Usage

perCycleBasecall(x)

## S4 method for signature 'RqcResultSet'
perCycleBasecall(x)

## S4 method for signature 'list'
perCycleBasecall(x)

perCycleQuality(x)

## S4 method for signature 'RqcResultSet'
perCycleQuality(x)

## S4 method for signature 'list'
perCycleQuality(x)

perFileInformation(x)

## S4 method for signature 'RqcResultSet'
perFileInformation(x)

## S4 method for signature 'list'
perFileInformation(x)

perFileTopReads(x)

## S4 method for signature 'RqcResultSet'
perFileTopReads(x)

## S4 method for signature 'list'
perFileTopReads(x)

perReadFrequency(x)

## S4 method for signature 'RqcResultSet'
perReadFrequency(x)

## S4 method for signature 'list'
perReadFrequency(x)

perReadQuality(x)

## S4 method for signature 'RqcResultSet'
perReadQuality(x)

## S4 method for signature 'list'
perReadQuality(x)

perReadWidth(x)

## S4 method for signature 'RqcResultSet'
perReadWidth(x)

## S4 method for signature 'list'
perReadWidth(x)

Arguments

x	RqcResultSet object or list of RqcResultSet objects

Value

data frame

data frame

data frame

data frame

data frame

data frame

data frame

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
head(perCycleBasecall(rqcResultSet))
head(perCycleQuality(rqcResultSet))
head(perReadFrequency(rqcResultSet))
head(perReadQuality(rqcResultSet))
head(perReadWidth(rqcResultSet))
perFileInformation(rqcResultSet)
perFileTopReads(rqcResultSet)

---

Interactive Quality Control Report

Description

This function runs a Shiny web application of interactive Rqc report. This is useful for large amount of files and sample groups.

Usage

rqcShinyReport(rqcResultSet)

Arguments

rqcResultSet

list of RqcResultSet-class objects

Value

function

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
# rqcShinyReport(rqcResultSet)

---

Minimun read mean quality and maximum percentage loss of reads estimations for trimming step.

Description

This function estimates how many reads would be lost if the sequences are filtered by a minimum read mean quality value. Also this function estimates what is the minimum read mean quality value for filtering and lose max percentage defined.

Usage

stats4trim(rqcResultSet, qmin, pmax)

Arguments

rqcResultSet	list of RqcResultSet objects created by rqc and rqcQA functions.
qmin	Minimum read mean quality value (bewteen 0 and 41).
pmax	Maximum percentage of reads permitted been lost during trimming step.

Value

A data frame containg estimated minimum quality and maximum percentage for each input file.

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
stats4trim(rqcResultSet, qmin=20)
stats4trim(rqcResultSet, pmax=10)

---

Subset RqcResultSet object by group name.

Description

This function subsets RqcResultSet object function by group name.

Usage

subsetByGroup(rqcResultSet, group)

Arguments

rqcResultSet	list of RqcResultSet objects created by rqc and rqcQA functions.
group	Name of the group to subset

Value

list of RqcResultSet objects from only one group.

Author(s)

Welliton Souza

Examples

folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
files <- list.files(full.names=TRUE, path=folder)
rqcResultSet <- rqcQA(files, workers=1, group=c("a", "b"))
perFileInformation(subsetByGroup(rqcResultSet, "a"))

---

Subset RqcResultSet object by pair files.

Description

This function subsets RqcResultSet object function by pair files.

Usage

subsetByPair(rqcResultSet, pair)

Arguments

rqcResultSet	list of RqcResultSet objects created by rqc and rqcQA functions.
pair	index of the pair

Value

list of RqcResultSet objects from only one pair.

Author(s)

Welliton Souza

Examples

checkpoint("Rqc", path=system.file(package="Rqc", "extdata"), {
  folder <- system.file(package="ShortRead", "extdata/E-MTAB-1147")
  files <- list.files(full.names=TRUE, path=folder)
  rqcResultSet <- rqcQA(files, pair=c(1,1), workers=1)
}, keep="rqcResultSet")
perFileInformation(subsetByPair(rqcResultSet, 1))

---

Title: Convert DNA sequences to Reduced Representation format

Description

This function receives a vector of strings (character vector) containing DNA sequences and returns a vector of strings containing codified DNA.

Usage

toRRDNA(dnas)

Arguments

dnas	Vector of DNA sequences (character vector).

Value

Vector of DNA converted to reduced representation format (character vector).

Note

This function is used internally to compute top over-represented reads and to store in RqcResultSet objects (per file top reads).

Author(s)

Welliton Souza

See Also

perFileTopReads

Examples

dna <- "ATCGNATCGTA"
dna.converted <- toRRDNA(dna)
nchar(dna)
nchar(dna.converted)

---