示例#1
0
def main(argv=None):
    """script main.

    parses command line options in sys.argv, unless *argv* is given.
    """

    if argv is None:
        argv = sys.argv

    # setup command line parser
    parser = U.OptionParser(version="%prog version: $Id$",
                            usage=globals()["__doc__"])

    parser.add_option("-p",
                      "--bc-pattern",
                      dest="pattern",
                      type="string",
                      help="Barcode pattern")
    parser.add_option("--bc-pattern2",
                      dest="pattern2",
                      type="string",
                      help="Barcode pattern for paired reads")
    parser.add_option("--3prime",
                      dest="prime3",
                      action="store_true",
                      help="barcode is on 3' end of read.")
    parser.add_option("--read2-in",
                      dest="read2_in",
                      type="string",
                      help="file name for read pairs")
    parser.add_option("--read2-out",
                      dest="read2_out",
                      type="string",
                      help="file to output processed paired read to")
    parser.add_option(
        "--read2-out-only",
        dest="read2_out_only",
        action="store_true",
        help="Paired reads, only output the second read in the pair")
    parser.add_option("--quality-filter-threshold",
                      dest="quality_filter_threshold",
                      type="int",
                      help=("Remove reads where any UMI base quality score "
                            "falls below this threshold"))
    parser.add_option(
        "--quality-filter-mask",
        dest="quality_filter_mask",
        type="int",
        help=("If a UMI base has a quality below this threshold, "
              "replace the base with 'N'"))
    parser.add_option("--quality-encoding",
                      dest="quality_encoding",
                      type="choice",
                      choices=["phred33", "phred64", "solexa"],
                      help=("Quality score encoding. Choose from 'phred33'"
                            "[33-77] 'phred64' [64-106] or 'solexa' [59-106]"))
    parser.add_option("--extract-method",
                      dest="extract_method",
                      type="choice",
                      choices=["string", "regex"],
                      help=("How to extract the umi +/- cell barcodes, Choose "
                            "from 'string' or 'regex'"))
    parser.add_option("--filter-cell-barcode",
                      dest="filter_cell_barcode",
                      action="store_true",
                      help="Filter the cell barcodes")
    parser.add_option("--error-correct-cell",
                      dest="error_correct_cell",
                      action="store_true",
                      help=("Correct errors in the cell barcode"))
    parser.add_option("--error-correct-threshold",
                      dest="error_correct_threshold",
                      type="int",
                      help=("Hamming distance allowed for correction"))
    parser.add_option("--plot-prefix",
                      dest="plot_prefix",
                      type="string",
                      help=("Prefix for plots to visualise the automated "
                            "detection of the number of 'true' cell barcodes"))
    parser.add_option("--output-whitelist",
                      dest="output_whitelist",
                      type="string",
                      help=("Write out the automatically generated whitelist"))
    parser.add_option("--whitelist-tsv",
                      dest="whitelist_tsv",
                      type="string",
                      help=("A whitelist of accepted cell barcodes"))
    parser.add_option("--blacklist-tsv",
                      dest="blacklist_tsv",
                      type="string",
                      help=("A blacklist of accepted cell barcodes"))
    parser.add_option(
        "--cell-barcode-subset",
        dest="cell_barcode_subset",
        type="int",
        help=("Use only the first N reads to automatically "
              "identify the true cell barcodes. If N is greater "
              "than the number of reads, all reads will be used"))
    parser.add_option("--reads-subset",
                      dest="reads_subset",
                      type="int",
                      help=("Only extract from the first N reads. If N is "
                            "greater than the number of reads, all reads will "
                            "be used"))
    parser.add_option(
        "--reconcile-pairs",
        dest="reconcile",
        action="store_true",
        help=("Allow the presences of reads in read2 input that are"
              "not present in read1 input. This allows cell barcode"
              "filtering of read1s without considering read2s"))
    parser.set_defaults(extract_method="string",
                        filter_cell_barcodes=False,
                        whitelist_tsv=None,
                        blacklist_tsv=None,
                        error_correct_cell=False,
                        error_correct_threshold=1,
                        pattern=None,
                        pattern2=None,
                        read2_in=None,
                        read2_out=False,
                        read2_out_only=False,
                        quality_filter_threshold=None,
                        quality_encoding=None,
                        plot_prefix=None,
                        output_whitelist=None,
                        cell_barcode_subset=50000000,
                        reconcile=False)

    # add common options (-h/--help, ...) and parse command line

    (options, args) = U.Start(parser, argv=argv)

    if options.quality_filter_threshold or options.quality_filter_mask:
        if not options.quality_encoding:
            U.error("must provide a quality encoding (--quality-"
                    "encoding) to filter UMIs by quality (--quality"
                    "-filter-threshold) or mask low quality bases "
                    "with (--quality-filter-mask)")

    if not options.pattern and not options.pattern2:
        if not options.read2_in:
            U.error("Must supply --bc-pattern for single-end")
        else:
            U.error("Must supply --bc-pattern and/or --bc-pattern "
                    "if paired-end ")

    if options.pattern2:
        if not options.read2_in:
            U.error("must specify a paired fastq ``--read2-in``")

        if not options.pattern2:
            options.pattern2 = options.pattern

    extract_cell = False
    extract_umi = False

    # If the pattern is a regex we can compile the regex(es) prior to
    # ExtractFilterAndUpdate instantiation
    if options.extract_method == "regex":
        if options.pattern:
            try:
                options.pattern = regex.compile(options.pattern)
            except regex.error:
                U.error("barcode_regex '%s' is not a "
                        "valid regex" % options.pattern)

        if options.pattern2:
            try:
                options.pattern2 = regex.compile(options.barcode_regex2)
            except regex.Error:
                U.error("barcode_regex2 '%s' is not a "
                        "valid regex" % options.barcode_regex2)

    # check whether the regex contains a umi group(s) and cell groups(s)
    if options.extract_method == "regex":
        if options.pattern:
            for group in options.pattern.groupindex:
                if group.startswith("cell_"):
                    extract_cell = True
                elif group.startswith("umi_"):
                    extract_umi = True
        if options.pattern2:
            for group in options.pattern2.groupindex:
                if group.startswith("cell_"):
                    extract_cell = True
                elif group.startswith("umi_"):
                    extract_umi = True

    # check whether the pattern string contains umi/cell bases
    elif options.extract_method == "string":
        if options.pattern:
            if "C" in options.pattern:
                extract_cell = True
            if "N" in options.pattern:
                extract_umi = True
        if options.pattern2:
            if "C" in options.pattern2:
                extract_cell = True
            if "N" in options.pattern2:
                extract_umi = True

    if options.whitelist_tsv:
        if options.blacklist_tsv:
            U.error("Do not supply a blacklist and a whitelist. Just "
                    "remove the blacklist barcodes from the whitelist!")

    if not extract_umi:
        if options.extract_method == "string":
            U.error("barcode pattern(s) do not include any umi bases "
                    "(marked with 'Ns') %s, %s" %
                    (options.pattern, options.pattern2))
        elif options.extract_method == "regex":
            U.error("barcode regex(es) do not include any umi groups "
                    "(starting with 'umi_') %s, %s" (options.pattern,
                                                     options.pattern2))

    if options.stdin == sys.stdin:
        if not options.whitelist_tsv and options.filter_cell_barcode:
            U.error(
                "cannot support reading from stdin if correcting cell barcode")
        read1s = umi_methods.fastqIterate(U.openFile(options.stdin))
    else:
        read1s = umi_methods.fastqIterate(U.openFile(options.stdin.name))

    # set up read extractor
    ReadExtractor = umi_methods.ExtractFilterAndUpdate(
        options.extract_method, options.pattern, options.pattern2,
        options.prime3, extract_cell, options.quality_encoding,
        options.quality_filter_threshold, options.quality_filter_mask,
        options.filter_cell_barcode)

    if options.filter_cell_barcode:
        if (not options.whitelist_tsv) or options.error_correct_cell:
            cell_barcode_counts = collections.Counter()

            n_reads = 0
            if not options.read2_in:
                for read1 in read1s:
                    n_reads += 1
                    cell_barcode = ReadExtractor.getCellBarcode(read1)
                    if cell_barcode:
                        cell_barcode_counts[cell_barcode] += 1
                    if options.cell_barcode_subset:
                        if (n_reads > options.cell_barcode_subset):
                            break
            else:
                read2s = umi_methods.fastqIterate(U.openFile(options.read2_in))
                for read1, read2 in izip(read1s, read2s):
                    n_reads += 1
                    cell_barcode = ReadExtractor.getCellBarcode(read1, read2)
                    if cell_barcode:
                        cell_barcode_counts[cell_barcode] += 1
                    if options.cell_barcode_subset:
                        if (n_reads > options.cell_barcode_subset):
                            break

            if options.blacklist_tsv:
                cell_blacklist = umi_methods.getUserDefinedBarcodes(
                    options.blacklist_tsv)
                for cell in cell_blacklist:
                    del cell_barcode_counts[cell]

            if options.whitelist_tsv:
                cell_whitelist = umi_methods.getUserDefinedBarcodes(
                    options.whitelist_tsv)
                error_correct_mappings = umi_methods.getErrorCorrectMappings(
                    cell_barcode_counts.keys(), cell_whitelist,
                    options.error_correct_threshold)
            else:
                # getCellWhitelist has not been properly defined yet!
                cell_whitelist, error_correct_mappings = umi_methods.getCellWhitelist(
                    cell_barcode_counts, options.error_correct_threshold,
                    options.plot_prefix)

            # re-make the reads1s iterator
            read1s = umi_methods.fastqIterate(U.openFile(options.stdin.name))

        else:
            cell_whitelist = umi_methods.getUserDefinedBarcodes(
                options.whitelist_tsv)
            error_correct_mappings = None, None

        false_to_true_map, true_to_false_map = error_correct_mappings

        if options.output_whitelist:

            with U.openFile(options.output_whitelist, "w") as outf:

                columns = [
                    "barcode", "count", "corrected_barcodes",
                    "corrected_barcode_counts"
                ]
                outf.write("\t".join(columns) + "\n")

                for barcode in sorted(list(cell_whitelist)):

                    if true_to_false_map:
                        corrected_barcodes = ",".join(
                            sorted(true_to_false_map[barcode]))
                        corrected_barcode_counts = ",".join(
                            map(str, [
                                cell_barcode_counts[x]
                                for x in sorted(true_to_false_map[barcode])
                            ]))
                    else:
                        corrected_barcodes, corrected_barcode_counts = "", ""

                    outf.write("%s\t%s\t%s\t%s\n" %
                               (barcode, cell_barcode_counts[barcode],
                                corrected_barcodes, corrected_barcode_counts))

        ReadExtractor.cell_whitelist = cell_whitelist
        ReadExtractor.false_to_true_map = false_to_true_map

    if options.read2_in is None:
        for read in read1s:
            new_read = ReadExtractor(read)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                        options.reads_subset):
                    break

            if not new_read:
                continue

            options.stdout.write(str(new_read) + "\n")

    else:
        read2s = umi_methods.fastqIterate(U.openFile(options.read2_in))

        if options.read2_out:
            read2_out = U.openFile(options.read2_out, "w")

        if options.reconcile:
            strict = False
        else:
            strict = True

        for read1, read2 in umi_methods.joinedFastqIterate(
                read1s, read2s, strict):
            reads = ReadExtractor(read1, read2)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                        options.reads_subset):
                    break

            if not reads:
                continue
            else:
                new_read1, new_read2 = reads

            if not options.read2_out_only:
                options.stdout.write(str(new_read1) + "\n")

            if options.read2_out:
                read2_out.write(str(new_read2) + "\n")

    if options.read2_out:
        read2_out.close()

    for k, v in ReadExtractor.getReadCounts().most_common():
        U.info("%s: %s" % (k, v))

    U.Stop()
示例#2
0
def main(argv=None):
    """script main.

    parses command line options in sys.argv, unless *argv* is given.
    """

    if argv is None:
        argv = sys.argv

    # setup command line parser
    parser = U.OptionParser(version="%prog version: $Id$",
                            usage=globals()["__doc__"])

    parser.add_option("-p", "--bc-pattern", dest="pattern", type="string",
                      help="Barcode pattern")
    parser.add_option("--bc-pattern2", dest="pattern2", type="string",
                      help="Barcode pattern for paired reads")
    parser.add_option("--3prime", dest="prime3", action="store_true",
                      help="barcode is on 3' end of read.")
    parser.add_option("--read2-in", dest="read2_in", type="string",
                      help="file name for read pairs")
    parser.add_option("--read2-out", dest="read2_out", type="string",
                      help="file to output processed paired read to")
    parser.add_option("--read2-stdout", dest="read2_stdout",
                      action="store_true",
                      help="Paired reads, send read2 to stdout, discarding read1")
    parser.add_option("--quality-filter-threshold",
                      dest="quality_filter_threshold", type="int",
                      help=("Remove reads where any UMI base quality score "
                            "falls below this threshold"))
    parser.add_option("--quality-filter-mask",
                      dest="quality_filter_mask", type="int",
                      help=("If a UMI base has a quality below this threshold, "
                            "replace the base with 'N'"))
    parser.add_option("--quality-encoding",
                      dest="quality_encoding", type="choice",
                      choices=["phred33", "phred64", "solexa"],
                      help=("Quality score encoding. Choose from 'phred33'"
                            "[33-77] 'phred64' [64-106] or 'solexa' [59-106]"))
    parser.add_option("--extract-method",
                      dest="extract_method", type="choice",
                      choices=["string", "regex"],
                      help=("How to extract the umi +/- cell barcodes, Choose "
                            "from 'string' or 'regex'"))
    parser.add_option("--filter-cell-barcode",
                      dest="filter_cell_barcode",
                      action="store_true",
                      help="Filter the cell barcodes")
    parser.add_option("--error-correct-cell",
                      dest="error_correct_cell",
                      action="store_true",
                      help=("Correct errors in the cell barcode"))
    parser.add_option("--whitelist",
                      dest="whitelist", type="string",
                      help=("A whitelist of accepted cell barcodes"))
    parser.add_option("--blacklist",
                      dest="blacklist", type="string",
                      help=("A blacklist of accepted cell barcodes"))
    parser.add_option("--reads-subset",
                      dest="reads_subset", type="int",
                      help=("Only extract from the first N reads. If N is "
                            "greater than the number of reads, all reads will "
                            "be used"))
    parser.add_option("--reconcile-pairs",
                      dest="reconcile", action="store_true",
                      help=("Allow the presences of reads in read2 input that are"
                            "not present in read1 input. This allows cell barcode"
                            "filtering of read1s without considering read2s"))
    parser.set_defaults(extract_method="string",
                        filter_cell_barcodes=False,
                        whitelist=None,
                        blacklist=None,
                        error_correct_cell=False,
                        pattern=None,
                        pattern2=None,
                        read2_in=None,
                        read2_out=False,
                        read2_stdout=False,
                        quality_filter_threshold=None,
                        quality_encoding=None,
                        reconcile=False)

    # add common options (-h/--help, ...) and parse command line

    (options, args) = U.Start(parser, argv=argv,
                              add_group_dedup_options=False,
                              add_sam_options=False)

    if options.quality_filter_threshold or options.quality_filter_mask:
        if not options.quality_encoding:
            U.error("must provide a quality encoding (--quality-"
                    "encoding) to filter UMIs by quality (--quality"
                    "-filter-threshold) or mask low quality bases "
                    "with (--quality-filter-mask)")

    if not options.pattern and not options.pattern2:
        if not options.read2_in:
            U.error("Must supply --bc-pattern for single-end")
        else:
            U.error("Must supply --bc-pattern and/or --bc-pattern "
                    "if paired-end ")

    if options.pattern2:
        if not options.read2_in:
            U.error("must specify a paired fastq ``--read2-in``")

        if not options.pattern2:
            options.pattern2 = options.pattern

    extract_cell = False
    extract_umi = False

    # If the pattern is a regex we can compile the regex(es) prior to
    # ExtractFilterAndUpdate instantiation
    if options.extract_method == "regex":
        if options.pattern:
            try:
                options.pattern = regex.compile(options.pattern)
            except regex.error:
                U.error("barcode_regex '%s' is not a "
                        "valid regex" % options.pattern)

        if options.pattern2:
            try:
                options.pattern2 = regex.compile(options.pattern2)
            except regex.Error:
                U.error("barcode_regex2 '%s' is not a "
                        "valid regex" % options.pattern2)

    # check whether the regex contains a umi group(s) and cell groups(s)
    if options.extract_method == "regex":
        if options.pattern:
            for group in options.pattern.groupindex:
                if group.startswith("cell_"):
                    extract_cell = True
                elif group.startswith("umi_"):
                    extract_umi = True
        if options.pattern2:
            for group in options.pattern2.groupindex:
                if group.startswith("cell_"):
                    extract_cell = True
                elif group.startswith("umi_"):
                    extract_umi = True

    # check whether the pattern string contains umi/cell bases
    elif options.extract_method == "string":
        if options.pattern:
            if "C" in options.pattern:
                extract_cell = True
            if "N" in options.pattern:
                extract_umi = True
        if options.pattern2:
            if "C" in options.pattern2:
                extract_cell = True
            if "N" in options.pattern2:
                extract_umi = True

    if not extract_umi:
        if options.extract_method == "string":
            U.error("barcode pattern(s) do not include any umi bases "
                    "(marked with 'Ns') %s, %s" % (
                        options.pattern, options.pattern2))
        elif options.extract_method == "regex":
            U.error("barcode regex(es) do not include any umi groups "
                    "(starting with 'umi_') %s, %s" (
                        options.pattern, options.pattern2))

    if options.filter_cell_barcodes:

        if not options.whitelist:
                U.error("must provide a whitelist (--whitelist) if using "
                        "--filter-cell-barcode option")

        if not extract_cell:
            if options.extract_method == "string":
                U.error("barcode pattern(s) do not include any cell bases "
                        "(marked with 'Cs') %s, %s" % (
                            options.pattern, options.pattern2))
            elif options.extract_method == "regex":
                U.error("barcode regex(es) do not include any cell groups "
                        "(starting with 'cell_') %s, %s" (
                            options.pattern, options.pattern2))

    read1s = umi_methods.fastqIterate(options.stdin)

    # set up read extractor
    ReadExtractor = umi_methods.ExtractFilterAndUpdate(
        options.extract_method,
        options.pattern,
        options.pattern2,
        options.prime3,
        extract_cell,
        options.quality_encoding,
        options.quality_filter_threshold,
        options.quality_filter_mask,
        options.filter_cell_barcode)

    if options.filter_cell_barcode:
        cell_whitelist, false_to_true_map = umi_methods.getUserDefinedBarcodes(
            options.whitelist, options.error_correct_cell)

        ReadExtractor.cell_whitelist = cell_whitelist
        ReadExtractor.false_to_true_map = false_to_true_map

    if options.blacklist:
        blacklist = set()
        with U.openFile(options.blacklist, "r") as inf:
            for line in inf:
                blacklist.add(line.strip().split("\t")[0])
        ReadExtractor.cell_blacklist = blacklist

    # variables for progress monitor
    progCount = 0
    displayMax = 100000
    U.info("Starting barcode extraction")

    if options.read2_in is None:
        for read in read1s:

            # incrementing count for monitoring progress
            progCount += 1

            # Update display in every 100kth iteration
            if progCount % displayMax == 0:
                U.info("Parsed {} reads".format(progCount))

            new_read = ReadExtractor(read)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                    options.reads_subset):
                    break

            if not new_read:
                continue

            options.stdout.write(str(new_read) + "\n")

    else:
        read2s = umi_methods.fastqIterate(U.openFile(options.read2_in))

        if options.read2_out:
            read2_out = U.openFile(options.read2_out, "w")

        if options.reconcile:
            strict = False
        else:
            strict = True

        for read1, read2 in umi_methods.joinedFastqIterate(
                read1s, read2s, strict):

            # incrementing count for monitoring progress
            progCount += 1

            # Update display in every 100kth iteration
            if progCount % displayMax == 0:
                U.info("Parsed {} reads".format(progCount))
                sys.stdout.flush()

            reads = ReadExtractor(read1, read2)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                    options.reads_subset):
                    break

            if not reads:
                continue
            else:
                new_read1, new_read2 = reads

            if options.read2_stdout:
                options.stdout.write(str(new_read2) + "\n")
            else:
                options.stdout.write(str(new_read1) + "\n")

                if options.read2_out:
                    read2_out.write(str(new_read2) + "\n")

    if options.read2_out:
        read2_out.close()

    for k, v in ReadExtractor.getReadCounts().most_common():
        U.info("%s: %s" % (k, v))

    U.Stop()
示例#3
0
def main(argv=None):
    """script main.

    parses command line options in sys.argv, unless *argv* is given.
    """

    if argv is None:
        argv = sys.argv

    # setup command line parser
    parser = U.OptionParser(version="%prog version: $Id$",
                            usage=globals()["__doc__"])

    group = U.OptionGroup(parser, "extract-specific options")

    # (Experimental option) Retain the UMI in the sequence read"
    group.add_option("--retain-umi",
                     dest="retain_umi",
                     action="store_true",
                     help=optparse.SUPPRESS_HELP)
    group.add_option("--read2-out",
                     dest="read2_out",
                     type="string",
                     help="file to output processed paired read to")
    group.add_option(
        "--read2-stdout",
        dest="read2_stdout",
        action="store_true",
        help="Paired reads, send read2 to stdout, discarding read1")
    group.add_option("--quality-filter-threshold",
                     dest="quality_filter_threshold",
                     type="int",
                     help=("Remove reads where any UMI base quality score "
                           "falls below this threshold"))
    group.add_option("--quality-filter-mask",
                     dest="quality_filter_mask",
                     type="int",
                     help=("If a UMI base has a quality below this threshold, "
                           "replace the base with 'N'"))
    group.add_option("--quality-encoding",
                     dest="quality_encoding",
                     type="choice",
                     choices=["phred33", "phred64", "solexa"],
                     help=("Quality score encoding. Choose from 'phred33'"
                           "[33-77] 'phred64' [64-106] or 'solexa' [59-106]"))
    group.add_option("--filter-cell-barcode",
                     dest="filter_cell_barcode",
                     action="store_true",
                     help="Filter the cell barcodes")
    group.add_option("--error-correct-cell",
                     dest="error_correct_cell",
                     action="store_true",
                     help=("Correct errors in the cell barcode"))
    group.add_option("--whitelist",
                     dest="whitelist",
                     type="string",
                     help=("A whitelist of accepted cell barcodes"))
    group.add_option("--blacklist",
                     dest="blacklist",
                     type="string",
                     help=("A blacklist of accepted cell barcodes"))
    group.add_option("--subset-reads",
                     "--reads-subset",
                     dest="reads_subset",
                     type="int",
                     help=("Only extract from the first N reads. If N is "
                           "greater than the number of reads, all reads will "
                           "be used"))
    group.add_option("--reconcile-pairs",
                     dest="reconcile",
                     action="store_true",
                     help=("Allow the presences of reads in read2 input that "
                           "are not present in read1 input. This allows cell "
                           "barcode filtering of read1s without "
                           "considering read2s"))

    parser.add_option_group(group)

    group = U.OptionGroup(parser, "[EXPERIMENTAl] barcode extraction options")

    group.add_option(
        "--either-read",
        dest="either_read",
        action="store_true",
        help="UMI may be on either read (see "
        "--either-read-resolve) for options to resolve cases where"
        "UMI is on both reads")
    group.add_option("--either-read-resolve",
                     dest="either_read_resolve",
                     type="choice",
                     choices=["discard", "quality"],
                     help=("How to resolve instances where both reads "
                           "contain a UMI but using --either-read."
                           "Choose from 'discard' or 'quality'"
                           "(use highest quality). default=dicard"))

    parser.add_option_group(group)

    parser.set_defaults(extract_method="string",
                        filter_cell_barcodes=False,
                        whitelist=None,
                        blacklist=None,
                        error_correct_cell=False,
                        pattern=None,
                        pattern2=None,
                        read2_in=None,
                        read2_out=False,
                        read2_stdout=False,
                        quality_filter_threshold=None,
                        quality_encoding=None,
                        reconcile=False,
                        either_read=False,
                        either_read_resolve="discard")

    # add common options (-h/--help, ...) and parse command line

    (options, args) = U.Start(parser,
                              argv=argv,
                              add_extract_options=True,
                              add_group_dedup_options=False,
                              add_umi_grouping_options=False,
                              add_sam_options=False)

    if options.retain_umi and not options.extract_method == "regex":
        raise ValueError(
            "option --retain-umi only works with --extract-method=regex")

    if options.quality_filter_threshold or options.quality_filter_mask:
        if not options.quality_encoding:
            U.error("must provide a quality encoding (--quality-"
                    "encoding) to filter UMIs by quality (--quality"
                    "-filter-threshold) or mask low quality bases "
                    "with (--quality-filter-mask)")

    extract_cell, extract_umi = U.validateExtractOptions(options)

    if options.either_read:
        if extract_cell:
            U.error("Option to extract from either read (--either-read) "
                    "is not currently compatible with cell barcode extraction")
        if not options.extract_method == "regex":
            U.error("Option to extract from either read (--either-read)"
                    "requires --extract-method=regex")
        if not options.pattern or not options.pattern2:
            U.error("Option to extract from either read (--either-read)"
                    "requires --bc-pattern=[PATTERN1] and"
                    "--bc-pattern2=[PATTERN2]")

    if options.filter_cell_barcodes:

        if not options.whitelist:
            U.error("must provide a whitelist (--whitelist) if using "
                    "--filter-cell-barcode option")

        if not extract_cell:
            if options.extract_method == "string":
                U.error("barcode pattern(s) do not include any cell bases "
                        "(marked with 'Cs') %s, %s" %
                        (options.pattern, options.pattern2))
            elif options.extract_method == "regex":
                U.error("barcode regex(es) do not include any cell groups "
                        "(starting with 'cell_') %s, %s" (options.pattern,
                                                          options.pattern2))

    read1s = umi_methods.fastqIterate(options.stdin)

    # set up read extractor
    ReadExtractor = umi_methods.ExtractFilterAndUpdate(
        options.extract_method, options.pattern, options.pattern2,
        options.prime3, extract_cell, options.quality_encoding,
        options.quality_filter_threshold, options.quality_filter_mask,
        options.filter_cell_barcode, options.retain_umi, options.either_read,
        options.either_read_resolve)

    if options.filter_cell_barcode:
        cell_whitelist, false_to_true_map = umi_methods.getUserDefinedBarcodes(
            options.whitelist, options.error_correct_cell)

        ReadExtractor.cell_whitelist = cell_whitelist
        ReadExtractor.false_to_true_map = false_to_true_map

    if options.blacklist:
        blacklist = set()
        with U.openFile(options.blacklist, "r") as inf:
            for line in inf:
                blacklist.add(line.strip().split("\t")[0])
        ReadExtractor.cell_blacklist = blacklist

    # variables for progress monitor
    progCount = 0
    displayMax = 100000
    U.info("Starting barcode extraction")

    if options.read2_in is None:
        for read in read1s:

            # incrementing count for monitoring progress
            progCount += 1

            # Update display in every 100kth iteration
            if progCount % displayMax == 0:
                U.info("Parsed {} reads".format(progCount))

            new_read = ReadExtractor(read)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                        options.reads_subset):
                    break

            if not new_read:
                continue

            options.stdout.write(str(new_read) + "\n")

    else:
        read2s = umi_methods.fastqIterate(U.openFile(options.read2_in))

        if options.read2_out:
            read2_out = U.openFile(options.read2_out, "w")

        if options.reconcile:
            strict = False
        else:
            strict = True

        for read1, read2 in umi_methods.joinedFastqIterate(
                read1s, read2s, strict):

            # incrementing count for monitoring progress
            progCount += 1

            # Update display in every 100kth iteration
            if progCount % displayMax == 0:
                U.info("Parsed {} reads".format(progCount))
                sys.stdout.flush()

            reads = ReadExtractor(read1, read2)

            if options.reads_subset:
                if (ReadExtractor.read_counts['Input Reads'] >
                        options.reads_subset):
                    break

            if not reads:
                continue
            else:
                new_read1, new_read2 = reads

            if options.read2_stdout:
                options.stdout.write(str(new_read2) + "\n")
            else:
                options.stdout.write(str(new_read1) + "\n")

                if options.read2_out:
                    read2_out.write(str(new_read2) + "\n")

    if options.read2_out:
        read2_out.close()

    for k, v in ReadExtractor.getReadCounts().most_common():
        U.info("%s: %s" % (k, v))

    U.Stop()