Getting Started

The short tutorial below explains how to run PROBer using a small example. Please go over the following steps before you continue:

  1. Download PROBer.
  2. Install PROBer.
  3. Read the manual.

If you want PROBer to align raw reads for you, you also need to install Bowtie and/or Bowtie 2.

Example

Suppose we have arabidopsis genome and gene annotation in two files: ‘TAIR10_chr_all.fa’ and ‘TAIR10_GFF3_genes.gff’. We choose the reference name as ‘arabidopsis’ and are only interested in mRNA and rRNA. The data we have are single-end reads with read length 37bp, with minus channel reads in ‘minus.fq’ and plus channel reads in ‘plus.fq’. The primer length is 6bp, the size selection range is from 21bp to 526bp. We use Bowtie aligner to align reads and assume Bowtie executables are under ‘/sw/bowtie’. We choose sample name as ‘test_sample’. We use 40 cores. In the end, we simulate 10M single-end reads with output name ‘test_sim’.

The commands are listed below:

PROBer prepare --gff3 TAIR10_GFF3_genes.gff --gff3-RNA-pattern mRNA,rRNA --bowtie --bowtie-path /sw/bowtie TAIR10_chr_all.fa arabidosis/arabidosis
PROBer estimate -p 40 --primer-length 6 --size-selection-min 21 --size-selection-max 526 --read-length 37 --bowtie-path /sw/bowtie arabidosis/arabidosis test_sample --reads plus.fq minus.fq
PROBer simulate arabidosis/arabidosis test_sample.temp/test_sample_minus.config test_sample minus 10000000 test_sim
PROBer simulate arabidosis/arabidosis test_sample.temp/test_sample_plus.config test_sample plus 10000000 test_sim