autoBLAST

A set of Python and Bash scripts for automated BLAST analysis of the flanking sequences of circular RNAs.

Installation

To install autoBLAST, run this in the terminal:

git clone https://github.com/alexandruioanvoda/autoBLAST
cd ./autoBLAST
chmod +x whole_intron_fasta.sh intron_fasta_window.sh master_script.sh blast\ RCM.py distance_analysis.py

The installation and runs were tested on OS X Yosemite and Ubuntu 16.10. Both computers had the following dependencies installed: Python 3.5.2, biopython 1.69, bedtools v2.26.0, Mac bash-3.2 or GNU bash 4.3 For any issues running scripts in this repository, please email us at avoda@nevada.unr.edu

Input

Two possible inputs: either for whole-introns (see IntronPicker @ https://github.com/alexandruioanvoda/IntronPicker) or fixed intervals flanking the splice sites (included in these scripts).

1. Whole introns:

If you identified flanking intron intervals by using IntronPicker, paste its output (both upstream_introns.bed & downstream_introns.bed) files into the autoBLAST folder

2. Fixed window size:

autoBLAST can run without the linked intron intervals, just specify the window intervals (in LL, LD, DL, DD, as in the diagram underneath) when running autoBLAST.

Running the analysis

Testing that it works

Open bash, navigate to the folder where you downloaded the scripts and run this command: get -c http://hgdownload.cse.ucsc.edu/goldenPath/ce11/bigZips/chromFa.tar.gz -O - | gzip -dc | tar -xO > ce11.fa This will download the ce11 genome FASTA file, which we'll use for extracting flanking intron sequences. After this, extract the test.zip file and run mv ./test/* ./. This will move all the BED6 files in the test folder into the autoBLAST folder. Now, run ./master_script.sh 1 test c.bed ce11.fa 400 0 0 400. This will BLAST and analyze 400bp seqs flanking the circRNAs to the left and to the right. The results should look the same as the ones included in the output.zip file.

Running your own samples

The first argument must be a 0 (whole-intron analysis) or 1 (flanking sized-window analysis). ./master_script.sh 0 samplename circRNAbed genomefastafile OR ./master_script.sh 1 samplename circRNAbed genomefastafile LL LD DL DD

Example: ./master_script.sh 1 WormsSeqFridayWindow400bpRCManalysis circRNA_intervals.bed ce11_genome.fa 400 0 0 400 OR ./master_script.sh 0 WormsSeqFridayWholeIntronRCM circRNA_intervals.bed ce11_genome.fa