Aligning reads to SILVA

As I was thinking about subsampling reads to make more fair comparisons I realized that there is probably still at least a small proportion of contaminating rRNA reads left in the transcriptomes. If I subsample before filtering out those reads, I may be losing out of data mapping to genes I care about downstream. In order to avoid this I decided to map the reads with Bowtie to the SILVA rRNA gene database and use only those reads that fail to find a match for anything later on in the pipeline.

I started by getting the non-redundant full length sequences from release 123 off of the mothur site. This file has sequences from bacteria (152308), eukaryotes (16209), and archea (3901). It looks like this:

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AB006051.LobTirol 
......................................................ATG---------CGTACGCCATGCGTG-------
GGCTAGACT------CGTAC------------------------------ACGTACGACACGTCCATAGACTCGATAC----------
...

In order to align reads to this file, I needed to remove the gaps before making the Bowtie database. I used the mothur command that removes gaps as follows:

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Degap.seqs(fasta=silva.nr_v123.align)

This gives you a fasta file with just sequence names and nucleic acids remaining, no more gaps. Making the bowtie database is almost as simple with the follow command:

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/home/mljenior/bin/bowtie/bowtie-build silva.nr_v123.ng.fasta silva_db

Now it’s ready to be used for filtering out rRNA reads from my metatranscriptomes. I’ll post the results from how well it does probably later this week.

Published Again!

After a long review process, the final publication from my dissertation work is finally published! In it were able to show that not only ...… Continue reading

Major Changes!

Published on August 17, 2017