Reference Code backup Executable files
It’s very important to check if current sequencing depth is deep enough to perform alternative splicing analyses. For a well annotated organism, the number of expressed genes in particular tissue is almost fixed so the number of splice junctions is also fixed. The fixed splice junctions can be predetermined from reference gene model. All (annotated) splice junctions should be rediscovered from a saturated RNA-seq data, otherwise, downstream alternative splicing analysis is problematic because low abundance splice junctions are missing. This module checks for saturation by resampling 5%, 10%, 15%, ..., 95% of total alignments from BAM or SAM file, and then detects splice junctions from each subset and compares them to reference gene model.
junction_saturation.py -i Pairend_nonStrandSpecific_36mer_Human_hg19.bam -r hg19.refseq.bed12 -o output
| --version | show program’s version number and exit |
| -h, --help | show this help message and exit |
| -i INPUT_FILE, --input-file=INPUT_FILE | |
| Alignment file in BAM or SAM format.[required] | |
| -o OUTPUT_PREFIX, --out-prefix=OUTPUT_PREFIX | |
| Prefix of output files(s). [required] | |
| -r REFGENE_BED, --refgene=REFGENE_BED | |
| Reference gene model in bed fomat. This gene model is used to determine known splicing junctions. [required] | |
| -l PERCENTILE_LOW_BOUND, --percentile-floor=PERCENTILE_LOW_BOUND | |
| Sampling starts from this percentile. A integer between 0 and 100. default=5 | |
| -u PERCENTILE_UP_BOUND, --percentile-ceiling=PERCENTILE_UP_BOUND | |
| Sampling ends at this percentile. A integer between 0 and 100. default=100 | |
| -s PERCENTILE_STEP, --percentile-step=PERCENTILE_STEP | |
| Sampling frequency. Smaller value means more sampling times. A integer between 0 and 100. default=5 | |
| -m MINIMUM_INTRON_SIZE, --min-intron=MINIMUM_INTRON_SIZE | |
| Minimum intron size (bp). default=50 | |
| -v MINIMUM_SPLICE_READ, --min-coverage=MINIMUM_SPLICE_READ | |
| Minimum number of supportting reads to call a junction. default=1 | |
| -q MAP_QUAL, --mapq=MAP_QUAL | |
| Minimum mapping quality (phred scaled) for an alignment to be called “uniquely mapped”. default=30 | |