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MarginPolish-HELEN walkthough (Local istall)

This document provides a guideline on how to use the polishing pipeline.

Local install walkthrough

This provides a guideline on how to install and run MP-HELEN locally.

Install MP-HELEN

Make sure you have all the prerequisites installed:

sudo apt-get -y install git cmake make gcc g++ autoconf bzip2 lzma-dev zlib1g-dev \
libcurl4-openssl-dev libpthread-stubs0-dev libbz2-dev liblzma-dev libhdf5-dev \
python3-pip python3-virtualenv virtualenv

Install HELEN:

git clone https://github.com/kishwarshafin/helen.git
cd helen
make install
. ./venv/bin/activate

helen --help
marginpolish --help
Setup input-output directory
mkdir mp_helen_walkthough
cd mp_helen_walkthough

mkdir mp_helen_models
mkdir mp_output
mkdir helen_output

export MODEL_OUTPUT_DIR="mp_helen_models"
export MP_OUTPUT_DIR="mp_output"
export HELEN_OUTPUT_DIR="helen_output"
Download data

The total download size is ~1.6GB.

wget https://storage.googleapis.com/kishwar-helen/bacterial_data/guppy_305/validation_data/Reads_to_assembly_StaphAur.bam
wget https://storage.googleapis.com/kishwar-helen/bacterial_data/guppy_305/validation_data/Reads_to_assembly_StaphAur.bam.bai
wget https://storage.googleapis.com/kishwar-helen/bacterial_data/guppy_305/validation_data/Draft_assembly_StaphAur.fasta
STEP 1: Download models

First download and save all the available models.

helen download_models \
--output_dir $MODEL_OUTPUT_DIR
STEP 2: Run MarginPolish

Next, run MarginPolish to generate images for HELEN.

marginpolish \
Reads_to_assembly_StaphAur.bam \
Draft_assembly_StaphAur.fasta \
$MODEL_OUTPUT_DIR/MP_r941_guppy344_microbial.json \
-t 30 \
-o $MP_OUTPUT_DIR/mp_images \
-f
STEP 3: Run HELEN

Finally, run HELEN to get the polished sequence:

helen polish \
--image_dir $MP_OUTPUT_DIR \
--model_path $MODEL_OUTPUT_DIR/HELEN_r941_guppy344_microbial.pkl \
--threads 38 \
--output_dir $HELEN_OUTPUT_DIR/ \
--output_prefix Staph_Aur_draft_helen

Add parameter --gpu if you have CUDA installed in your machine. You can now exit the venv by simply typing exit.

exit

Assessment of the assembly (Optional)

You can assess the assembly using Pomoxis. Make sure you exit the venv of HELEN.

Install Pomoxis

sudo apt-get install cmake wget bzip2 zlib1g-dev libncurses5-dev \
python3-all-dev libhdf5-dev libatlas-base-dev libopenblas-base \
libopenblas-dev libbz2-dev liblzma-dev libffi-dev make python-virtualenv
git clone --recursive https://github.com/nanoporetech/pomoxis
cd pomoxis
make install
. ./venv/bin/activate

Download reference quality assembly

wget https://storage.googleapis.com/kishwar-helen/bacterial_data/guppy_305/validation_data/truth_assembly_staph_aur.fasta

Run assess_assembly

mkdir pomoxis_assessment
cd pomoxis_assessment

assess_assembly \
-i ../Draft_assembly_StaphAur.fasta \
-r ../truth_assembly_staph_aur.fasta \
-p draft_assembly_quality \
-l 50 \
-t 32 \
-e \
-T

Expected Output:

#  Q Scores
  name     mean      q10      q50      q90
 err_ont  24.16      inf    24.59    22.74
 err_bal  24.15      inf    24.58    22.71
    iden  31.61      inf    32.66    30.85
     del  32.10      inf    34.61    31.46
     ins  25.95      inf    26.38    23.00

Now assess the polished assembly:

assess_assembly \
-i ../helen_output/Staph_Aur_draft_helen.fa \
-r ../truth_assembly_staph_aur.fasta \
-p polished_assembly_quality \
-l 50 \
-t 32 \
-e \
-T

Expected output:

#  Q Scores
  name     mean      q10      q50      q90
 err_ont  33.90    35.09    33.98    32.84
 err_bal  33.90    35.09    33.98    32.84
    iden  38.88    40.97    39.19    37.21
     del  39.19    42.44    39.21    37.59
     ins  38.03    40.09    37.45    36.93