A tool for the analysis of amplicon methylation data. Works with the output from Bismark, a program used to map bisulfite treated sequencing reads to the genome of interest and perform methylation calls.
The tool delivers the following:
- Classify the genomic sequences by their methylation status into: mostly methylated, mostly unmethylated and partially methylated. Methylation status refers to the number of CpGs in a genomic sequence that are methylated
- Phase the genomic sequences according to a heterozygous SNP in the amplicon region
- Count the number of genomic sequences in each methylation status class and report it per group of phased reads
Options:
| Option | Type | Description |
|---|---|---|
| --inpath | PATH | Directory with CpG and alignment files files [required] |
| --thr | FLOAT | Threshold for allele frequency bellow which an allele is ignored [required] |
| --outpath | PATH | Path to place the output [required] |
| --ampltable | PATH | Tab separated file with amplicon attributes [required] |
| --plotgrid | string | Number of plots to draw per row and column, separated with ";" |
-
--inpath a directory is provided where the output the alignment files and the methylation calls is placed. Script will first try to match files with glob (bam), extract the sample name and then look for the methylation call files (CpG). All file names are expected to have the format as output by bismark aligner and bismark_methylation_extractor
-
--ampltable a tab separated file is provided, with the fields as shown in the following example:
| Name | Chr | start | end | strand | size_bp | upper_mCG_thr | low_mCG_thr | snps_coord |
|---|---|---|---|---|---|---|---|---|
| H19 | chr11 | 1999754 | 2000063 | neg | 310 | 20 | 3 | 1999845;1999934 |
Most of the fields are self explanatory. The rest are as:
- upper_mCG_CG is an integer. If a read has this number, or more, CpGs methylated, it is classified as methylated.
- low_mCG_thr is an integer. If a read has this number, or less, CpGs methylated, it is classified as unmethylated.
- snps_coord is an integer used to provide the location(s) of the heterozygous SNP(s) on which the reads are phased. If more than one, they should be separated with ;. If no SNPs are present, a - must be placed in that field.