StringFix: an annotation-guided transcriptome assembler

Brief introduction

• StringFix recover transcript sequence and the corresponding amino acid sequence by taking small variants (INDEL and SNP) into account.
• It optionally utilizes a previous annotations (in GTF or GFF file format) to generate amino acid sequences while reflecting small variants found in the alignment of reads to a reference genome.
• You can use either the GTF annotation associated with the reference genome or the GFF file generated by some reference-based transcriptome assmeblers, such as StringTie, Cufflink, Strawberry and so on.
• Basically, the SW recovers amino acid sequence from the transcript sequence based on the annotations in GTF/GFF, if there is no changes in start/stop codon.
• While, if start/stop codon is garbled, it searches a new open reading frame (ORF) by applying the 'longest ORF rule'.
• cite: Lee J, Kim M, Han K, and Yoon S, StringFix: an annotation-guided transcriptome assembler improves the recovery of amino acid sequences from RNA-Seq reads, Genes & Genimics https://doi.org/10.1007/s13258-023-01458-7

Installation using pip

StringFix can be installed using pip command. With python3 installed in your system. Use the follwing command in a terminal.

pip install StringFix

StringFix requires the pre-installed python packages including Numpy, Pandas, sklearn, and scipy.

Command line usage of StringFix

• run_sfix.py is a python script that can be run in a terminal.

• Download run_sfix.py to your local directory and simply run the script by typing python3 run_sfix.py or python run_sfix.py in a terminal to see the available parameters.

• According to what information is provided, StringFix is run in one of the following modes of operation.

  1. Genome-guided mode: requires only SAM or BAM file (a mandatory parameter) (pybam is used to load BAM file.)
  2. Annotation-guided mode: requires SAM/BAM and GTF/GFF annotation, where, for the latter, one can use either the reference GTF associated to the reference genome or the GFF file generated by some genome-guided transcriptome assemblers
  3. Annotation-guided mode with sequence correction: requires SAM/BAM, GTF/GFF annotation and the reference genome (The reference genome must be the one used for read alignment)

• Once it is successfully run, it outputs the following files

  1. transcriptome.fasta, containing the transcript sequences for all the isoforms that were detected as 'expressed' with its estimated coverage above the threshold
  2. Expression profiles in 'tsv' format, containing the raw read coverage depth and TPM for all the transcript detected
  3. SNV summary in 'tsv' format, containg all the small variations, including indel and SNP. SNPs are provided only if the path to the reference genome is provided
  4. proteome.fasta, containing the amino acid sequences for all the protein-coding genes that were detected as 'expressed' with its estimated coverage above the threshold. (Path to the reference genome must be given to get protein sequences)

Example data and script

1. Click here to download the test data and store it to your preferred directory.
2. decompress the file by typing tar -xzvf StringFix_test_data.tar.gz in a terminal.
3. Change directory to the decompressed directory.
4. run ./test_run.sh. (If needed, make the shell script executable by typing chmod 777 test_run.sh)

Contact

Send email to syoon@dku.edu for any inquiry on the usages.