How to align reads to known ncRNA database

skblazer · 12-29-2010, 01:17 PM

I want to align a lot of reads to the known ncRNA database.

I don't know which mapper is fit for small RNA alignment.

blast? megablast? or bowtie ...

how to set the parameter?

Thanks

colindaven · 01-04-2011, 05:35 AM

You'll need to mention what read length and system you used for the sequencing. Generally BLAST is too slow for "a lot of reads".

dcfargo · 01-04-2011, 06:02 AM

DSAP aligns to both miRBase (16) and RFam (10) if you have very short read data

http://dsap.cgu.edu.tw/

skblazer · 01-04-2011, 09:59 AM

Thanks

My question is if I used blast/megablast, I'll get a lot of alignments without perfect aligning rate. How to set the cutoff to filter the output?

For example, if one read's 6-26bp (27bp in total) can be aligned to a known tRNA, is it a candidate tRNA or not?

For tRNA or rRNA, I should use a more relax alignment than miRNA, am I right?

bioinfosm · 01-04-2011, 04:36 PM

another option could be to align all reads to the genome without biasing to a selective reference dataset, and then use coordinates of your ncRNA database of interest, and identify which ones are expressed?

skblazer · 01-05-2011, 08:18 AM

Because the organism I'm working on does not have a reference, I have to align them to some ncRNA database.

I don't know the if the similarity of tRNA or rRNA among the organims is high as well as miRNA.

Quote:

Originally Posted by bioinfosm

another option could be to align all reads to the genome without biasing to a selective reference dataset, and then use coordinates of your ncRNA database of interest, and identify which ones are expressed?

colindaven · 01-06-2011, 12:27 AM

To the best of my knowledge rRNA and tRNAs are _more_ conserved than miRNAs. At least in bacteria we work on there is considerable variation in miRNAs between closely related species.
I'm sure you can find some good references on this in pubmed though.

http://www.biomedcentral.com/1471-2164/8/481
"Our results suggest that while there is a conserved set of miRNAs among plant species, a large fraction of miRNAs vary among species"

Ultimately you'll need to consider topics like thermodynamic stability, perhaps this link is helpful ?
http://www.biomedcentral.com/1471-2148/10/329/abstract

sdarko · 01-06-2011, 05:26 AM

Quote:

Originally Posted by skblazer

Thanks

My question is if I used blast/megablast, I'll get a lot of alignments without perfect aligning rate. How to set the cutoff to filter the output?

For example, if one read's 6-26bp (27bp in total) can be aligned to a known tRNA, is it a candidate tRNA or not?

For tRNA or rRNA, I should use a more relax alignment than miRNA, am I right?

I've been using BLASTn to align snRNA reads to tRNA, rRNA, snoRNA, piwi associated RNA and miRNA.

Basically, I first trim 3' adapter sequence and collapse the reads. Then I use BLASTn and use '-perc_identity 100' and '-word_size 16' so that at least 16 bases have to perfectly align to get a hit. When I'm parsing through my results I compare the length of the transcript and the length of the alignment. If they're the same, I call it a good alignment. If they're not, I set it aside to align to my next reference set.

skblazer · 01-07-2011, 05:32 PM

Thanks for your advise.

I'll try.

Similar Threads
Thread	Thread Starter	Forum	Replies	Last Post
Tophat for finding long ncRNA with short reads?	KevinLam	Bioinformatics	3	02-24-2017 10:11 AM
Align genomic DNA sequence to protein database?	rdu	Bioinformatics	0	11-03-2011 08:31 PM
Reads that can align to multiple places	ashwatha	Bioinformatics	2	09-21-2011 08:21 PM
Align reads to contigs	ojy	Bioinformatics	3	07-25-2011 10:16 AM
map solid reads to ncRNA database	crh	SOLiD	2	07-19-2010 12:22 PM

12-29-2010, 01:17 PM	#1
skblazer Member Location: Massachusetts Join Date: Feb 2009 Posts: 50	How to align reads to known ncRNA database I want to align a lot of reads to the known ncRNA database. I don't know which mapper is fit for small RNA alignment. blast? megablast? or bowtie ... how to set the parameter? Thanks

01-04-2011, 04:36 PM	#5
bioinfosm Senior Member Location: USA Join Date: Jan 2008 Posts: 482	another option could be to align all reads to the genome without biasing to a selective reference dataset, and then use coordinates of your ncRNA database of interest, and identify which ones are expressed? __________________ -- bioinfosm

01-04-2011, 05:35 AM	#2
colindaven Senior Member Location: Germany Join Date: Oct 2008 Posts: 415	You'll need to mention what read length and system you used for the sequencing. Generally BLAST is too slow for "a lot of reads".

01-04-2011, 06:02 AM	#3
dcfargo Member Location: Chapel Hill Join Date: Aug 2008 Posts: 22	DSAP aligns to both miRBase (16) and RFam (10) if you have very short read data http://dsap.cgu.edu.tw/

01-04-2011, 09:59 AM	#4
skblazer Member Location: Massachusetts Join Date: Feb 2009 Posts: 50	Thanks My question is if I used blast/megablast, I'll get a lot of alignments without perfect aligning rate. How to set the cutoff to filter the output? For example, if one read's 6-26bp (27bp in total) can be aligned to a known tRNA, is it a candidate tRNA or not? For tRNA or rRNA, I should use a more relax alignment than miRNA, am I right?

01-06-2011, 12:27 AM	#7
colindaven Senior Member Location: Germany Join Date: Oct 2008 Posts: 415	To the best of my knowledge rRNA and tRNAs are _more_ conserved than miRNAs. At least in bacteria we work on there is considerable variation in miRNAs between closely related species. I'm sure you can find some good references on this in pubmed though. http://www.biomedcentral.com/1471-2164/8/481 "Our results suggest that while there is a conserved set of miRNAs among plant species, a large fraction of miRNAs vary among species" Ultimately you'll need to consider topics like thermodynamic stability, perhaps this link is helpful ? http://www.biomedcentral.com/1471-2148/10/329/abstract

01-07-2011, 05:32 PM	#9
skblazer Member Location: Massachusetts Join Date: Feb 2009 Posts: 50	Thanks for your advise. I'll try.