Pipeline for phasiRNA detection

Question

[v.baaskarla]

Hi guys,

My question is regarding the pipeline for identification of phasiRNAs in plants.

There are many papers that have used an analysis method but were not very informative on how to execute the protocol.

Links to papers:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4457045/

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1867363/

http://link.springer.com/article/10.1007/s12042-016-9173-4

Method:

The PHAS loci and phasiRNAs were predicted as described previously (Zheng et al. 2014; Srivastava et al. 2015). The unique sequences in the small RNA libraries were mapped to the genome of pineapple with SOAP2 (Li et al. 2009). A self-developed program was used to scan the genome and cDNA sequences using a window of 210 nt or 240 nt (ten 21 nt or 24 nt) respectively. A two-nucleotide positive offset was used to calculate the positions of siRNAs on the anti-sense strand because the existence of two-nucleotide over-hang at the 3’-end of siRNA duplex (Howell et al. 2007; Xia et al. 2013; Chen et al. 2007; Zhai et al. 2011). Then a P-value was calculated for each of the windows using a modified version of methods in Chen et al. (2007),

I would like to know which program can be used to scan the genome with a specific window length as described (Bold) above.

I'd greatly appreciate inputs from anyone who has previous experience in small RNA and specifically phasiRNA analysis.

Thanks

V

Answer 1

[rosenkranz]

There are currently 2 published tools that can predict phasiRNAs. First one is PhaseTank, which runs on Linux and uses bowtie1 for mapping small RNA sequences to a reference. PhaseTank searches for phasiRNAs using the method described in the papers you listed above. The 210 and 240 nt sliding window size comes from the implemented algorithm which obligatory uses windows of 10 so-called cycle-bins (corresponds to the length of a phasiRNA). You will not be able to change that unless you recode the PhaseTank Perl script.

Software: http://phasetank.sourceforge.net/
Paper: https://academic.oup.com/bioinformatics/article-lookup/doi/10.1093/bioinformatics/btu628

Second tool is unitas (platform independent Perl script), which uses a SAM map file (you can use any aligner) as input. unitas uses a different algorithm for phasiRNA prediction which was found to be more sensitive without producing more false positive hits.

Software: http://www.smallrnagroup.uni-mainz.de/software.html
Paper: https://bmcgenomics.biomedcentral.com/articles/10.1186/s12864-017-4031-9

In contrast to PhaseTank, unitas is a universal annotation software. If you are interested in phasiRNAs only and you want to skip all the other annotation stuff, your terminal command could look like this:

perl unitas.pl -input map.file -species genus_species -skip_mapping -phasi 21 -phasi 24

Answer 2

[sqz986536]

Hi, I am finding the pipeline for phasiRNA detection. There is another software: PHASIS.