R/search.R

Defines functions filter_duplicates parsable_html parse_attributes get_options get_defaults get_refresh_from_meta parse_pre parse_a parse_primer_hits retry BLAST_primer print_options enumerate_ambiguity enumerate_primers primer_search

Documented in primer_search

#PrimerTree
#Copyright (C) 2013 Jim Hester

#' Query a pair of primers using ncbi's Primer-BLAST, if primers contain iupac
#'
#' ambiguity codes, enumerate all possible combinations and combine the
#' results.
#' @param forward forward primer to search by 5'-3' on plus strand
#' @param reverse reverse primer to search by 5'-3' on minus strand
#' @param num_aligns number of alignment results to keep
#' @param num_permutations the number of primer permutations to search, if the degenerate bases
#'        cause more than this number of permutations to exist, this number will be
#'        sampled from all possible permutations.
#' @param ... additional arguments passed to Primer-Blast
#' @param .parallel if 'TRUE', perform in parallel, using parallel backend
#'        provided by foreach
#' @param .progress name of the progress bar to use, see 'create_progress_bar'
#' @return data.frame of primer hits
#' @export
primer_search = function(forward, reverse, num_aligns=500, num_permutations=25, ..., .parallel=FALSE, .progress='none'){
  if(missing(forward) || missing(reverse))
    BLAST_primer()
  primers = enumerate_primers(forward, reverse)
  num_primers = nrow(primers)
  if(num_primers > num_permutations){
    warning(immediate.=T, 'primers have ', num_primers, ' possible combinations due to degenerate bases, sampling ', num_permutations, " primers, use 'num_permutations' to change")
    primers = primers[ sample.int(num_primers, num_permutations, replace=F), ]
  }
  message('BLASTing ', nrow(primers), ' primer combinations')
  #enumerate all combinations to handle ambiguity codes
  alply(primers, .margins=1, .expand=F, .parallel=.parallel, .progress=.progress,
        function(row) BLAST_primer(row$forward, row$reverse, num_targets_with_primers=max(num_aligns %/% nrow(primers), 1), ...))
}
iupac = list( "M" = list("A", "C"),
              "R" = list("A", "G"),
              "W" = list("A", "T"),
              "S" = list("C", "G"),
              "Y" = list("C", "T"),
              "K" = list("G", "T"),
              "V" = list("A", "C", "G"),
              "H" = list("A", "C", "T"),
              "D" = list("A", "G", "T"),
              "B" = list("C", "G", "T"),
              "N" = list("A", "C", "G", "T"),
              "I" = list("A", "T", "C"))

enumerate_primers = function(forward, reverse){
  forward_primers = enumerate_ambiguity(forward)
  data.frame(forward=forward_primers,
             reverse=rep(enumerate_ambiguity(reverse),
                         each=length(forward_primers)))
}
enumerate_ambiguity = function(sequence){
  search_regex = paste(names(iupac), collapse='|')
  locs = str_locate_all(sequence, search_regex)
  sequences = list()
  count = 1
  for (i in seq_len(nrow(locs[[1]]))){
    loc = locs[[1]][i,]
    ambiguity = str_sub(sequence, loc[1], loc[2])
    for(type in iupac[[ambiguity]]){
      new_seq = sequence
      str_sub(new_seq, loc[1], loc[2]) <- type
      sequences[[count]] = enumerate_ambiguity(new_seq)
      count = count + 1
    }
    return(unlist(sequences))
  }
  return(sequence)
}

print_options = function(options){
  output = capture.output( print(
                                 options[ is.na(options$type) | options$type != 'hidden',
                                        c('name', 'type', 'defval') ])
                          )

  message(paste(output, "\n", sep=""))
}

BLAST_primer = function(forward, reverse, ..., organism='',
  primer_specificity_database='nt', exclude_env='on'){

  url = 'http://www.ncbi.nlm.nih.gov/tools/primer-blast/'
  form = GET_retry(url)

  content = parsable_html(form)

  all_options = get_options(content)

  if(missing(forward) || missing(reverse)){
    print_options(all_options)
    stop('No primers specified')
  }

  options = list(..., primer_left_input=forward, primer_right_input=reverse,
                 organism=organism,
                 primer_specificity_database=primer_specificity_database,
                 exclude_env=exclude_env,
                 search_specific_primer='on')

  names(options) = toupper(names(options))

  match_args = pmatch(names(options), all_options$name)
  bad_args = is.na(match_args)

  if(any(bad_args)){
    print_options(all_options)
    stop(paste(names(options)[bad_args], collapse=','), ' not valid option\n')
  }

  options = get_defaults(options, all_options)

  start_time = now()

  message('Submitting Primer-BLAST query')
  response = POST_retry(paste(url, 'primertool.cgi', sep=''), body=options)

  values = get_refresh_from_meta(response)

  while(length(values) > 0){
    message('BLAST alignment processing, refreshing in ', values[1], ' seconds...')
    Sys.sleep(values[1])
    response = GET_retry(values[2])

    values = get_refresh_from_meta(response)
  }
  total_time = start_time %--% now()
  message('BLAST alignment completed in ', total_time %/% seconds(1), ' seconds')
  response
}

#modify a function to check the status and retry until success
retry = function(fun, num_retry=5, ...){
  function(...){
    res = fun(...)
    itr = 0
    status = http_status(res)
    while(itr < num_retry && inherits(res, 'response') && status$category != "success"){
      warning('request failed, retry attempt ', itr+1)
      res = fun(...)
      status = http_status(res)
      itr = itr + 1
    }
    res
  }
}

GET_retry = retry(GET)
POST_retry = retry(POST)

parse_primer_hits = function(response){
  content = parsable_html(response)
  rbind.fill(xpathApply(content, '//pre', parse_pre))
}
parse_a = function(a){
  #links like entrez/viewer.fcgi?db=nucleotide&id=452085006
  m = regexpr('id=\\d+', xmlAttrs(a)['href'])
  gi = gsub('id=', '', unlist(regmatches(xmlAttrs(a)['href'], m)))
  if(length(gi) == 0L){
    #links like nucleotide/449036831?from=1107741&to=1107929&report=gbwithparts
    m = regexpr('nucleotide/\\d+', xmlAttrs(a)['href'])
    gi = gsub('nucleotide/', '', unlist(regmatches(xmlAttrs(a)['href'], m)))
  }
  if(length(gi) == 0L) gi = NA
  data.frame(gi=as.character(gi), accession=as.character(xmlValue(a)))
}

parse_pre = function(pre){
  pre_text = xmlValue(pre)

  a = getNodeSet(pre, './preceding-sibling::a[1]')
  if(length(a) <= 0)
    stop('Parsing failed for ', pre_text)

  ids = parse_a(a[[1]])

  product_length_regex = 'product length = (\\d+)'
  template_regex = 'Template[^\\d]+(\\d+)[^.ACGT]+([.ACGT]+)[^\\d]+(\\d+)'
  full_regex = paste('[\\S\\W]*', product_length_regex, '[\\S\\W]*?',
                     template_regex, '[\\S\\W]*', template_regex, '[\\S\\W]*', sep='')
  values = str_split(gsub(full_regex, paste('\\', 1:8, sep='', collapse='|'),
                           pre_text, perl=T), '[|]')[[1]]
  data.frame(ids,
             product_length=as.numeric(values[1]),
             mismatch_forward=str_count(values[3], '[ACGT]'),
             mismatch_reverse=str_count(values[6], '[ACGT]'),
             forward_start = as.numeric(values[2]),
             forward_stop = as.numeric(values[4]),
             reverse_start = as.numeric(values[5]),
             reverse_stop = as.numeric(values[7]),
             product_start=min(as.numeric(values[c(2,4,5,7)])),
             product_stop=max(as.numeric(values[c(2,4,5,7)]))
             )
}
get_refresh_from_meta = function(response){
  content = parsable_html(response)
  meta = content['//meta[@http-equiv="Refresh"]']
  if(length(meta) > 0){
    values = str_split(xmlAttrs(meta[[1]])['content'], '; URL=')[[1]]
    return(values)
  }
  return()
}

get_defaults = function(set_options, options){
  #only look at values with set defaults
  options = options[ !is.na(options$defval), ]
  unchanged_options = setdiff(options$name, names(set_options))
  default_values = as.character(options[ match(unchanged_options, options$name), 'defval' ])
  names(default_values) = unchanged_options
  c(set_options, default_values)
}

get_options = function(content){
  options = rbind.fill(xpathApply(content, '//form//input | //form//select', parse_attributes))
  options$type = as.character(options$type)

  #add dropdown type if they are NA
  options$type[is.na(options$type)] <- 'dropdown'

  #make default values for checkboxes on or off rather than checked or unchecked
  options$defval = as.character(options$defval)
  check_map = c('checked' = 'on', 'unchecked' = '')
  checkboxes = which(options$type == 'checkbox')
  options$defval[ checkboxes ] = check_map[ options$defval[ checkboxes ] ]

  options[ options$type != 'hidden',  c('name', 'type', 'defval') ]
}

parse_attributes = function(x){
  as.data.frame(t(xmlAttrs(x)))
}
parsable_html = function(response){
  txt <- content(response, as='text')

  # remove any unicode characters
  Encoding(txt) <- "UTF-8"
  txt <- iconv(txt, "UTF-8", "ASCII", sub = "")

  #this gsub regex is to remove the definition lines, some of which have
  #  bracketed <junk> in them, which messes up the parsing
  txt <- gsub('("new_entrez".*?</a>).*?<pre>\n\n', '\\1\n<pre>', txt)
  htmlParse(txt)
}
filter_duplicates = function(hits){
  location_columns = c('accession', 'forward_start', 'forward_stop', 'reverse_start', 'reverse_stop')
  hits[!duplicated(t(apply(hits[location_columns], 1, range))),]
}

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primerTree documentation built on May 30, 2017, 4:15 a.m.