Create a script that executes the pipeline

.gitignore

@@ -1 +0,0 @@
-*.fastq

README.org

@@ -1,49 +1,3 @@
 * locigenesis
 
 locigenesis is a tool that generates an immune repertoire and runs it through a sequence reader simulation tool, to generate sequencing errors.
-
-** Installation
-
-This project uses [[https://nixos.org/][Nix]] to ensure reproducible builds.
-
-1. Install Nix (compatible with MacOS, Linux and [[https://docs.microsoft.com/en-us/windows/wsl/about][WSL]]):
-
-#+begin_src shell
-curl -L https://nixos.org/nix/install | sh
-#+end_src
-
-1. Clone the repository:
-
-#+begin_src shell
-git clone https://git.coolneng.duckdns.org/coolneng/locigenesis
-#+end_src
-
-3. Change the working directory to the project:
-
-#+begin_src shell
-cd locigenesis
-#+end_src
-
-4. Enter the nix-shell:
-
-#+begin_src shell
-nix-shell
-#+end_src
-
-After running these commands, you will find yourself in a shell that contains all the needed dependencies.
-
-** Usage
-
-An execution script that accepts 2 parameters is provided, the following command invokes it:
-
-#+begin_src shell
-./generation.sh <number of sequences> <number of reads>
-#+end_src
-
-- <number of sequences>: an integer that specifies the number of different sequences to generate
-- <number of reads>: an integer that specifies the number of reads to perform on each sequence
-
-The script will generate 2 files under the data directory:
-
-| HVR.fastq         | Contains the original CDR3 sequence                             |
-| CuReSim-HVR.fastq | Contains CDR3 after the read simulation, with sequencing errors |

docs/notebook.org

@@ -1,46 +0,0 @@
-#+TITLE: locigenesis
-#+AUTHOR: Amin Kasrou Aouam
-#+DATE: 2021-03-10
-* Sequence alignment
-
-Our generated sequences contain the full VJ region, but we are only interested in the CDR3 (Complementarity-determining region). We will proceed by delimiting CDR3, using the known sequences of V and J.
-
-#+begin_src R :results value silent
-v_segments <- readRDS("data/v_segments.rds")
-j_segments <- readRDS("data/j_segments_phe.rds")
-#+end_src
-
-#+begin_src R
-print(v_segments)
-print(j_segments)
-#+end_src
-
-#+RESULTS:
-#+begin_example
-  A DNAStringSet instance of length 147
-      width seq                                             names
-  [1]   326 GATACTGGAATTACCCAGACAC...ATCTCTGCACCAGCAGCCAAGA TRBV1*01_P
-  [2]   326 GATGCTGAAATCACCCAGAGCC...ATTTCTGCGCCAGCAGTGAGTC TRBV10-1*01_F
-  [3]   326 GATGCTGAAATCACCCAGAGCC...ATTTCTGCGCCAGCAGTGAGTC TRBV10-1*02_F
-  [4]   326 GATGCTGGAATCACCCAGAGCC...ATTTCTGCGCCAGCAGTGAGTC TRBV10-2*01_F
-  [5]   326 GATGCTGGAATCACCCAGAGCC...ATTTCTGCGCCAGCAGTGAGTC TRBV10-2*02_F
-  ...   ... ...
-[143]   324 GATACTGGAGTCTCCCAGAACC...GTATCTCTGTGCCAGCACGTTG TRBV7-9*06_(F)
-[144]   323 .........................TGTATCTCTGTGCCAGCAGCAG TRBV7-9*07_(F)
-[145]   325 GATTCTGGAGTCACACAAACCC...TATTTCTGTGCCAGCAGCGTAG TRBV9*01_F
-[146]   325 GATTCTGGAGTCACACAAACCC...TATTTCTGTGCCAGCAGCGTAG TRBV9*02_F
-[147]   321 GATTCTGGAGTCACACAAACCC...TTTGTATTTCTGTGCCAGCAGC TRBV9*03_(F)
-  A DNAStringSet instance of length 16
-     width seq                                              names
- [1]    32 TGGGCGTCTGGGCGGAGGACTCCTGGTTCTGG                 TRBJ2-2P*01_ORF
- [2]    31 TTTGGAGAGGGAAGTTGGCTCACTGTTGTAG                  TRBJ1-3*01_F
- [3]    31 TTTGGTGATGGGACTCGACTCTCCATCCTAG                  TRBJ1-5*01_F
- [4]    31 TTTGGCAGTGGAACCCAGCTCTCTGTCTTGG                  TRBJ1-4*01_F
- [5]    31 TTCGGTTCGGGGACCAGGTTAACCGTTGTAG                  TRBJ1-2*01_F
- ...   ... ...
-[12]    31 TTTGGCCCAGGCACCCGGCTGACAGTGCTCG                  TRBJ2-3*01_F
-[13]    31 TTCGGGCCAGGCACGCGGCTCCTGGTGCTCG                  TRBJ2-5*01_F
-[14]    31 TTCGGGCCAGGGACACGGCTCACCGTGCTAG                  TRBJ2-1*01_F
-[15]    31 TTCGGGCCGGGCACCAGGCTCACGGTCACAG                  TRBJ2-7*01_F
-[16]    31 GTCGGGCCGGGCACCAGGCTCACGGTCACAG                  TRBJ2-7*02_ORF
-#+end_example

generation.sh

@@ -1,22 +1,21 @@
 #!/bin/sh
 
 usage() {
-	echo "usage: generation.sh <number of sequences> <number of reads>"
+	echo "usage: generation.sh <number of sequences>"
 	exit 1
 }
 
-if [ $# != 2 ]; then
+if [ $# != 1 ]; then
 	usage
 fi
 
 sequences=$1
-number_of_reads=$2
 data_directory="data/"
-fastq=".fastq"
-filename="sequence"
 prefix="curesim_"
 
-Rscript src/repertoire.r "$sequences" "$number_of_reads" &&
+Rscript src/repertoire.r "$sequences"
-	CuReSim -f "$data_directory$filename$fastq" -o "$data_directory$prefix$filename$fastq"
+
-Rscript src/alignment.r
+for file in "$data_directory"*.fastq; do
-rm "$data_directory/log.txt"
+	file_name=$(cut -d / -f 2)
+	java -jar tools/CuReSim.jar -f "$file" -o "$data_directory$prefix$file_name"
+done

shell.nix

@@ -2,35 +2,14 @@
 
 with pkgs;
 
-let
+mkShell {
-  CuReSim = stdenv.mkDerivation rec {
-    name = "CuReSim";
-    version = "1.3";
-    src = fetchzip {
-      url =
-        "http://www.pegase-biosciences.com/wp-content/uploads/2015/08/${name}${version}.zip";
-      sha256 = "1hvlpgy4haqgqq52mkxhcl9i1fx67kgwi6f1mijvqzk0xff77hkp";
-      stripRoot = true;
-      extraPostFetch = ''
-        chmod go-w $out
-      '';
-    };
-
-    nativeBuildInputs = [ makeWrapper ];
-
-    installPhase = ''
-      mkdir -pv $out/share/java $out/bin
-      cp -r ${src} $out/share/java/${name}
-      makeWrapper ${pkgs.jdk}/bin/java $out/bin/CuReSim --add-flags "-jar $out/share/java/${name}/${name}.jar"
-    '';
-  };
-in mkShell {
   buildInputs = [
     R
     rPackages.immuneSIM
     rPackages.Biostrings
-    rPackages.stringr
     jdk
-    CuReSim
+    # Develoment tools
+    rPackages.languageserver
+    rPackages.lintr
   ];
 }

src/alignment.r

@@ -1,101 +0,0 @@
-library(Biostrings)
-library(parallel)
-
-parse_data <- function(file) {
-  reversed_sequences <- Biostrings::readQualityScaledDNAStringSet(file)
-  sequences <- Biostrings::reverseComplement(reversed_sequences)
-  vj_segments <- union(
-    readRDS("data/v_segments.rds"),
-    readRDS("data/j_segments_phe.rds")
-  )
-  return(list(sequences, vj_segments))
-}
-
-parse_metadata <- function(metadata) {
-  id_elements <- unlist(strsplit(metadata, split = " "))
-  v_identifier <- id_elements[2]
-  j_identifier <- id_elements[3]
-  return(list(v_id = v_identifier, j_id = j_identifier))
-}
-
-match_id_sequence <- function(names, vdj_segments, id) {
-  matches <- grep(names, pattern = id)
-  row <- matches[1]
-  return(as.character(vdj_segments[row]))
-}
-
-get_vj_sequence <- function(metadata, names, vdj_segments) {
-  identifiers <- parse_metadata(metadata)
-  v_sequence <- match_id_sequence(names, vdj_segments, id = identifiers["v_id"])
-  j_sequence <- match_id_sequence(names, vdj_segments, id = identifiers["j_id"])
-  return(list(v_seq = v_sequence, j_seq = j_sequence))
-}
-
-fetch_vj_sequences <- function(sequences, vdj_segments) {
-  vj_sequences <- sapply(names(sequences),
-    names(vdj_segments),
-    vdj_segments,
-    FUN = get_vj_sequence
-  )
-  results <- data.frame(t(vj_sequences))
-  return(results)
-}
-
-align_sequence <- function(sequence, vdj_segment) {
-  return(Biostrings::pairwiseAlignment(
-    subject = sequence,
-    pattern = vdj_segment,
-    type = "global-local",
-    gapOpening = 1
-  ))
-}
-
-handle_indels <- function(insertion, deletion, cys, alignment) {
-  ins_start <- sum(Biostrings::width(deletion[start(deletion) <= cys$start]))
-  ins_end <- sum(Biostrings::width(deletion[end(deletion) <= cys$end]))
-  shift_num <- c(0, cumsum(Biostrings::width(insertion))[-length(ins_start)])
-  shifted_ins <- IRanges::shift(insertion, shift_num)
-  gaps <- sum(width(shifted_ins[end(shifted_ins) < cys$start + ins_start])) +
-    nchar(stringr::str_extract(alignedSubject(alignment), "^-*"))
-  return(list("start" = ins_start - gaps, "end" = ins_end - gaps))
-}
-
-get_cys_coordinates <- function(alignment) {
-  cys <- list("start" = 310, "end" = 312)
-  insertion <- unlist(Biostrings::insertion(alignment))
-  deletion <- unlist(Biostrings::deletion(alignment))
-  delta_coordinates <- handle_indels(insertion, deletion, cys, alignment)
-  cys_start <- cys$start + delta_coordinates$start
-  cys_end <- cys$end + delta_coordinates$end
-  return(list("start" = cys_start, "end" = cys_end))
-}
-
-get_hvr_sequences <- function(sequences, vdj_segments, cores = detectCores()) {
-  df <- fetch_vj_sequences(sequences, vdj_segments)
-  v_alignment <- parallel::mcmapply(sequences,
-    df$v_seq,
-    FUN = align_sequence,
-    mc.cores = cores
-  )
-  cys_coordinates <- parallel::mclapply(v_alignment, FUN = get_cys_coordinates)
-  cys_df <- as.data.frame(do.call(rbind, cys_coordinates))
-  remaining <- Biostrings::subseq(sequences, start = unlist(cys_df$end))
-  j_alignment <- parallel::mcmapply(remaining,
-    df$j_seq,
-    FUN = align_sequence,
-    mc.cores = cores
-  )
-  j_start <- parallel::mclapply(
-    j_alignment,
-    function(x) start(Biostrings::Views(x)),
-    mc.cores = cores
-  )
-  hvr_start <- unlist(cys_df$start)
-  hvr_end <- unlist(cys_df$start) + unlist(j_start) + 2
-  hvr <- Biostrings::subseq(sequences, start = hvr_start, end = hvr_end)
-  return(hvr)
-}
-
-data <- parse_data(file = "data/curesim_sequence.fastq")
-hvr <- get_hvr_sequences(sequences = data[[1]], vdj_segments = data[[2]])
-Biostrings::writeXStringSet(hvr, "data/CuReSim-HVR.fastq", format = "fastq")

src/repertoire.r

@@ -1,42 +1,55 @@
 library(immuneSIM)
 library(Biostrings)
 
-generate_repertoire <- function(number_of_sequences) {
+generate_repertoires <- function(number_of_sequences) {
-  return(immuneSIM(
+  a_chain <- immuneSIM(
     number_of_seqs = number_of_sequences,
     species = "hs",
     receptor = "tr",
-    chain = "b"
+    chain = "a",
-  ))
+    verbose = TRUE
-}
-
-save_data <- function(data) {
-  Biostrings::writeXStringSet(data$sequence,
-    "data/sequence.fastq",
-    format = "fastq"
   )
-  Biostrings::writeXStringSet(data$junction, "data/HVR.fastq", format = "fastq")
+  b_chain <- immuneSIM(
+    number_of_seqs = number_of_sequences,
+    species = "hs",
+    receptor = "tr",
+    chain = "b",
+    verbose = TRUE
+  )
+  return(list("a_chain" = a_chain, "b_chain" = b_chain))
 }
 
-process_data <- function(data, reads) {
+process_chain <- function(repertoire) {
-  dna_sequence <- Biostrings::DNAStringSet(data$sequence)
+  sequences <- as.character(repertoire$sequence)
-  data$sequence <- Biostrings::reverseComplement(dna_sequence)
+  counts <- as.integer(repertoire$counts)
-  names(data$sequence) <- paste(rownames(data), data$v_call, data$j_call, " ")
+  reads <- Biostrings::DNAStringSet(rep(sequences, counts))
-  data$junction <- Biostrings::DNAStringSet(data$junction)
+  names(reads) <- seq_len(length(reads))
-  names(data$junction) <- rownames(data)
+  reverse_complement <- Biostrings::reverseComplement(reads)
-  amplified_data <- data[rep(seq_len(nrow(data)), reads), ]
+  return(reverse_complement)
-  return(amplified_data)
+}
+
+preprocess_data <- function(repertoires) {
+  filtered_repertoires <- lapply(repertoires, process_chain)
+  names(filtered_repertoires) <- names(repertoires)
+  return(filtered_repertoires)
+}
+
+save_data <- function(repertoires) {
+  for (chain in names(repertoires)) {
+    file_name <- paste("data/", chain, ".fastq", sep = "")
+    Biostrings::writeXStringSet(repertoires[[chain]], file_name, format = "fastq")
+  }
+}
+
+parse_cli_arguments <- function(args) {
+  if (length(args) != 1) {
+    stop("usage: repertoire.r <number of sequences>")
+  }
+  return(as.integer(args[1]))
 }
 
-parse_cli_arguments <- function() {
 args <- commandArgs(trailingOnly = TRUE)
-  if (length(args) != 2) {
+number_of_sequences <- parse_cli_arguments(args)
-    stop("usage: repertoire.r <number of sequences> <number of reads>")
+sim_repertoire <- generate_repertoires(number_of_sequences)
-  }
+processed_data <- preprocess_data(sim_repertoire)
-  return(args)
+save_data(processed_data)
-}
-
-args <- parse_cli_arguments()
-repertoire <- generate_repertoire(number_of_sequences = as.integer(args[1]))
-data <- process_data(data = repertoire, reads = args[2])
-save_data(data)