Sex Check Module

The usage of the standalone module is shown below:

$ pyGenClean_sex_check --help
usage: pyGenClean_sex_check [-h] [-v] --bfile FILE [--femaleF FLOAT]
                            [--maleF FLOAT] [--nbChr23 INT] [--gender-plot]
                            [--sex-chr-intensities FILE]
                            [--gender-plot-format FORMAT] [--lrr-baf]
                            [--lrr-baf-raw-dir DIR] [--lrr-baf-format FORMAT]
                            [--out FILE]

Check sample's gender using Plink.

optional arguments:
  -h, --help            show this help message and exit
  -v, --version         show program's version number and exit

Input File:
  --bfile FILE          The input file prefix (will find the Plink binary
                        files by appending the prefix to the .bed, .bim, and
                        .fam files, respectively.

Options:
  --femaleF FLOAT       The female F threshold. [default: < 0.300000]
  --maleF FLOAT         The male F threshold. [default: > 0.700000]
  --nbChr23 INT         The minimum number of markers on chromosome 23 before
                        computing Plink's sex check [default: 50]

Gender Plot:
  --gender-plot         Create the gender plot (summarized chr Y intensities
                        in function of summarized chr X intensities) for
                        problematic samples.
  --sex-chr-intensities FILE
                        A file containing alleles intensities for each of the
                        markers located on the X and Y chromosome for the
                        gender plot.
  --gender-plot-format FORMAT
                        The output file format for the gender plot (png, ps,
                        pdf, or X11 formats are available). [default: png]

LRR and BAF Plot:
  --lrr-baf             Create the LRR and BAF plot for problematic samples
  --lrr-baf-raw-dir DIR
                        Directory or list of directories containing
                        information about every samples (BAF and LRR).
  --lrr-baf-format FORMAT
                        The output file format for the LRR and BAF plot (png,
                        ps, pdf, or X11 formats are available). [default: png]

Output File:
  --out FILE            The prefix of the output files (which will be a Plink
                        binary file. [default: sexcheck]

Input Files

This module uses PLINK’s binary file format (bed, bim and fam files) for the source data set (the data of interest).

If the option of generating the gender plot is used, a file containing intensities information about each markers on the sexual chromosomes is required. This file (which could be gzipped) should contain at least the following columns:

• Sample ID: the unique sample id for each individual.
• SNP Name: the unique name of each markers.
• Chr: the name of the chromosome on which each marker is located.
• X: the intensities of the first allele of each marker.
• Y: the intensities of the second allele of each marker.

If the options of generating the BAF and LRR values is used, the name of a directory containing intensities file for each sample is required. The name of each file should be the unique sample id. This file (which could be gzipped) should contain at least the following columns:

• Chr: the name of the chromosome on which each marker is located.
• Position: the position of the marker on the chromosome.
• B Allele Freq: the BAF value of each marker.
• Log R Ratio: the LRR value of each marker.

If the two plotting module is used alone, one more file is required per module: a list of samples with gender problem and explanation for pyGenClean.SexCheck.gender_plot, and only the list of samples with gender problem for pyGenClean.SexCheck.baf_lrr_plot (both files are provided by the pyGenClean.SexCheck.sex_check module).

Procedure

Here are the steps performed by the module:

1. Checks if there are enough markers on the chromosome 23. If not, the module stops here.
2. Runs the sex check analysis using Plink.
3. If there are no sex problems, the module quits.
4. Creates the recoded file for the chromosome 23.
5. Computes the heterozygosity percentage on the chromosome 23.
6. If there are enough markers on chromosome 24 (at least 1), creates the recoded file for this chromosome.
7. Computes the number of no call on the chromosome 24.
8. If required, plots the gender plot.
   1. If there are summarized_intensities provided, reads the files and skips to step vi.
   2. Reads the bim file to get markers on the sexual chromosomes.
   3. Reads the fam file to get individual’s gender.
   4. Reads the file containing samples with sex problems.
   5. Reads the intensities and summarizes them.
   6. Plots the summarized intensities.
9. If required, plots the BAF and LRR plots.
   1. Reads the problematic samples.
   2. Finds and checks the raw files for each of the problematic samples.
   3. Plots the BAF and LRR plots.

Output Files

The output files of each of the steps described above are as follow (note that the output prefix shown is the one by default [i.e sexcheck]):

1. No output files created.
2. One set of PLINK’s result files:
   • sexcheck: the result of the sex check procedure from Plink.
3. Two files are created if there are sex problems:
   • sexcheck.list_problem_sex: a summary of samples with sex problem.
   • sexcheck.list_problem_sex_ids: the list of sample ids with sex problem.
4. One set of Plink’s files:
   • sexcheck.chr23_recodeA: the recoded file for the chromosome 23.
5. One custom output file:
   • sexcheck.chr23_recodeA.raw.hetero: the heterozygosity percentage on the chromosome 23. The file includes the following columns: PED (the pedigree ID), ID (the individual ID), SEX (the gender) and HETERO (the heterozygosity).
6. One set of Plink’s files:
   • sexcheck.chr24_recodeA: the recoded file for the chromosome 24.
7. One custom output file:
   • sexcheck.chr24_recodeA.raw.noCall: the number of no call on the chromosome 24. The file includes the following columns: PED (the pedigree ID), ID (the individual ID), SEX (the gender), nbGeno (the number of genotypes on the chromosome 24) and nbNoCall (the number of genotypes that were not called on chromosome 24).
8. Multiple files and one plot. The files are created so that the plot can be generated again with different parameters (since the summarized intensities for each sample are really long to compute).
   • sexcheck.png: the gender plot (see Figure Gender plot).
   • sexcheck.ok_females.txt: the list of females without sex problem, including their summarized intensities on chromosome 23 and 24.
   • sexcheck.ok_males.txt: the list of males without sex problem, including their summarized intensities on chromosome 23 and 24.
   • sexcheck.ok_unknowns.txt: the list of unknown gender without sex problem, including their summarized intensities on chromosome 23 and 24.
   • sexcheck.problematic_females.txt: the list of females with sex problem, including their summarized intensities on chromosome 23 and 24.
   • sexcheck.problematic_males.txt: the list of males with sex problem, including their summarized intensities on chromosome 23 and 24.
   • sexcheck.problematic_unknowns.txt: the list of unknown gender with sex problem, including their summarized intensities on chromosome 23 and 24. When this file is created by the pyGenClean.SexCheck.sex_check module, it is empty.
9. A directory containing one file per individual with gender problem (see Figure BAF and LRR plot).

The Plots

The plot generated by the pyGenClean.SexCheck.gender_plot module (the Gender plot figure) represents the summarized intensities of each sample of the data set. The color code represent the gender of each sample. Blue and red dots represent the males and females without gender problem, respectively. The green and purple triangles represent the males and females with gender problem. The gray dots and triangles represent the samples with unknown gender, with and without gender problem, respectively. This plot makes possible to find samples with sexual chromosomes abnormalities, such as males which are XXY or females with only one copy of the X chromosome (X0). Males that appear to be females and vice versa might in fact be sample mix up and would require further analysis.

Gender plot

The Gender plot figure can also be manually created after the data clean up pipeline, using its results and this following standalone script:

$ pyGenClean_gender_plot --help
usage: pyGenClean_gender_plot [-h] [-v] --bfile FILE [--intensities FILE]
                              [--summarized-intensities FILE] --sex-problems
                              FILE [--format FORMAT] [--xlabel STRING]
                              [--ylabel STRING] [--out FILE]

Plots the gender using X and Y chromosomes' intensities

optional arguments:
  -h, --help            show this help message and exit
  -v, --version         show program's version number and exit

Input File:
  --bfile FILE          The plink binary file containing information about
                        markers and individuals. Must be specified if
                        '--summarized-intensities' is not.
  --intensities FILE    A file containing alleles intensities for each of the
                        markers located on the X and Y chromosome. Must be
                        specified if '--summarized-intensities' is not.
  --summarized-intensities FILE
                        The prefix of six files (prefix.ok_females.txt,
                        prefix.ok_males.txt, prefix.ok_unknowns.txt,
                        problematic_females.txt, problematic_males.txt and
                        problematic_unknowns.txt) containing summarized chr23
                        and chr24 intensities. Must be specified if '--bfile'
                        and '--intensities' are not.
  --sex-problems FILE   The file containing individuals with sex problems.
                        This file is not read if the option 'summarized-
                        intensities' is used.

Options:
  --format FORMAT       The output file format (png, ps, pdf, or X11 formats
                        are available). [default: png]
  --xlabel STRING       The label of the X axis. [default: X intensity]
  --ylabel STRING       The label of the Y axis. [default: Y intensity]

Output File:
  --out FILE            The prefix of the output files (which will be a Plink
                        binary file. [default: sexcheck]

The log R ratio (LRR) for a sample is the log ratio of the normalized R value for the marker divided by the expected normalized R value. Hence, a value of 0 means 2 copies. A drop in the LRR shows a loss of a copy, while an increasing LRR shows a gain of a copy. Expected LRR values on chromosome 23 for female and female are 0 and [-0.5, -1], respectively. The LRR values of each markers on both the X and Y chromosomes are shown in the BAF and LRR plot figure.

The B allele frequency (BAF) for a sample shows the theta value for a marker, corrected for cluster position. For a normal number of copies, markers should have a BAF around 1 (homozygous for the B allele), 0.5 (heterozygous) or 0 (homozygous for A allele). Normal females should have the three lines across the chromosome. Normal males should only have two lines, located near 1 or 0. The BAF values of each markers on both the X and Y chromosomes are shown in the BAF and LRR plot figure.

BAF and LRR plot

The BAF and LRR plot figure can also be manually created after the data clean up pipeline, using this following standalone script:

$ pyGenClean_baf_lrr_plot --help
usage: pyGenClean_baf_lrr_plot [-h] [-v] --problematic-samples FILE
                               [--use-full-ids] [--full-ids-delimiter CHAR]
                               --raw-dir DIR [--format FORMAT] [--out FILE]

Plots the BAF and LRR of problematic samples.

optional arguments:
  -h, --help            show this help message and exit
  -v, --version         show program's version number and exit

Input File:
  --problematic-samples FILE
                        A file containing the list of samples with sex
                        problems (family and individual ID required, separated
                        by a single tabulation). Uses only the individual ID
                        by default, unless --use-full-ids is used.
  --use-full-ids        Use this options to use full sample IDs (famID and
                        indID). Otherwise, only the indID will be use.
  --full-ids-delimiter CHAR
                        The delimiter between famID and indID for the raw file
                        names. [default: _]
  --raw-dir DIR         Directory containing information about every samples
                        (BAF and LRR).

Options:
  --format FORMAT       The output file format (png, ps, pdf, or X11 formats
                        are available). [default: png]

Output File:
  --out FILE            The prefix of the output files. [default: sexcheck]

The Algorithm

For more information about the actual algorithms and source codes, refer to the following pages.

• pyGenClean.SexCheck.sex_check
• pyGenClean.SexCheck.gender_plot
• pyGenClean.SexCheck.baf_lrr_plot