excellent and well researched answer
What is the difference between non-coding RNAs and microRNAs. A protocol that I am following for predicting the miRNAs in a genome has a step in it where ncRNAs are filtered but I isn't miRNAs non coding RNAs in the first place? So why is this filter being implemented.
Reference
http://www.sciencedirect.com/science/article/pii/S0888754311001315
2 answers
miRNAs are indeed small non-coding RNAs. We can detect them by sequencing all RNAs in a given size range, map them to the corresponding genome, and look for the places where reads stack up in a pattern indicating that it's a miRNA precursor (basically: two stacks of reads with a given spacing, precise drop-off in read coverage that indicates precise Dicer processing, the resulting sequence folds into a stable hairpin)
That pattern may, however, also be found at sites generating other types of small structural RNAs, e.g. tRNAs. Removing all sites/sequences known to generating other types of structural RNAs is an efficient heuristic that will remove a lot of "false positives" that we would otherwise have to identify as "non-miRNAs" by close inspection of the data, or might miss altogether.
In a recent study where we detected miRNAs in a non-model organism (but with a sequenced genome), we did it as follows:
Abundant noncoding RNAs (rRNAs, tRNAs, snRNAs, and snoRNAs) were annotated to the N. vectensis genome using BLASTN (version 2.2.18) to identify homologs of
- Drosophila melanogaster annotated rRNAs, tRNAs, snRNAs, and snoRNAs (downloaded from FlyBase and GenBank),
- the most abundant rRNA variants found in previous studies involving sequencing of small RNAs (Ghildiyal et al. 2008; Seitz et al. 2008),
- Homo sapiens rRNAs, snRNAs, and snoRNAs (downloaded from GenBank, and the LBME human snoRNA database), and
- the complete set of tRNAs found in the Genomic tRNA database.
Genomic sequences with a significant BLAST hit (E-value < 10^-4) to any of those databases were flagged as "abundant noncoding RNAs," and deep-sequencing reads showing perfect identity to one of those were discarded.
MicroRNAs are a type of non-coding RNAs. Other types of ncRNA are for example snoRNAs, siRNAs, snRNAs, exRNAs, and piRNAs and long ncRNAs. I guess that the filtering steps would be based on size or sequence motif.
How do you suggest that I weed out miRNA from the rest of the non-coding RNAs. The paper which I have linked says that it was done with Blastn but I just cant understand how that would suffice.
They probably blast all the obtained sequences and when they are already known in the database they remove them I guess.
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