secondary_rewriter

Some aligners such as minimap2 do not write the SEQ and QUAL fields to secondary alignments (https://github.com/lh3/minimap2/issues/458) making it hard to analyze them (for example, SNPs will not be visible in a genome browser for secondary alignments). This program adds these back, referring to the primary alignment to get the SEQ and QUAL, and adding them to the secondaries. This is a two- (or three- for sorted output) pass program. The first pass collects the secondary alignments and the second adds the SEQ and QUAL fields to the secondaries.

Install

First install rust, probably with rustup https://rustup.rs/

Then

cargo install secondary_rewriter

Usage

```

First pass

samtools view yourfile.bam | secondary_rewriter --pass1 > secondaries.txt

Second pass

samtools view -h yourfile.bam | secondary_rewriter --pass2 --secondaries secondaries.txt | samtools view -o out.bam

Third pass, sort the BAM since all the new secondaries are added at the end of the file

samtools sort out.bam -o out.sorted.bam ```

You can package this into a small bash script

```

!/bin/bash

write_secondaries.sh

example usage

./write_secondaries.sh input.cram ref.fa output.cram

samtools view -@3 $1 -T $2 | secondaryrewriter --pass1 > secondaries.txt samtools view -@3 -h $1 -T $2 | secondaryrewriter --pass2 --secondaries secondaries.txt | samtools view -@3 -T $2 - -o $3 ```

The two-pass strategy works as follows

  1. First pass: output ALL secondary alignments (reads with flag 256) to a separate file
  2. Second pass: reading secondary alignments into memory, and then scan original SAM/BAM/CRAM to add SEQ and QUAL fields encountered during scan to the secondary alignments that are stored in a hashmap

It uses a two-pass strategy because otherwise it would have to effectively load the entire SAM/BAM/CRAM into memory

Help

```

% secondaryrewriter --help secondaryrewriter 0.1.0 Adds SEQ and QUAL fields to secondary alignments which are often missing from minimap2

USAGE: secondary_rewriter [OPTIONS]

OPTIONS: -h, --help Print help information --output-only-new-data --pass1 --pass2 -s, --secondaries -V, --version Print version information

```

--output-only-new-data only outputs the secondary alignments with their new SEQ/QUAL fields (and skips all other data). the default mode without this flag passes all other alignments through stdout and adds the secondary alignments at the end

Note

My second rust project!

```

```