long_read_polishing

Assembly Polishing using Racon

Racon is a GPU-accelerated polishing tool for assembly from long reads.

Installation

Installation instructions for Racon are available at https://github.com/lbcb-sci/racon#installation.

Data

Racon requires 3 files to generate a polished assembly -

1. Read data (in .fasta or .fastq for, with or without compression)
2. Overlap information of reads to reference (in .paf, .sam or mhap)
3. Target/reference sequences (in .fasta or .fastq for, with or without compression)

Oxford Nanopore read for SARS-CoV-2 can be downloaded from https://sra-pub-src-1.s3.amazonaws.com/SRR10948550/HKU-SZ-002a.fastq.1

Reference sequence for the virus is available here.

Usage

The following commands demostrate how to run a reference guided assembly using long read data and polishing. The example requires minimap2 to be available as well (https://github.com/lh3/minimap2).

minimap2 -x map-ont -t 12 MN908947.fasta HKU-SZ-002a.fastq > overlaps.paf
racon -c 4 -m 8 -x -6 -g -8 -w 500 -t 12 -q -1 HKU-SZ-002a.fastq overlaps.paf MN908947.fasta > assembled.fasta

Output of racon is a polished assembly. The following is an excerpt from a polishing run -

$ head -n6 assembled.fasta
>MN908947 LN:i:29910 RC:i:1000 XC:f:1.000000
ATTAAAGGTTTATACCTTCCCAGGTAACAAACCAACCAACTTTCGATCTCTTGTAGATCTGTTCTCTAAACGAACTTTA
AAATCTGTGTGGCTGTCACTCGGCTGCATGCTTAGTGCACTCACGCGGTATAATTAATAACTAATTACTGTCGTTGACA
GGACACGAGTAACTCGTCTATCTTCTGCAGGCTGCTTACGGTTTCGTCCGTGTTGCAGCCGATCATCAACACATCTAGG
TTTCGTCCGGGTGTGACCGAAAGGTAAGATGGAGAGCCTTGTCCCTGGTTTCAACGAGAAAACACACGTCCAACTCAGT
TTGCCTGTTTTACAGGTTCGCGACGTGCTCGTACGTGGCTTTGGAGACTCCGTGGAGGAGGTCTTATCAGAGGCACGTC