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Using miR-PREFeR: is this a microRNA?

mir-prefer mirdeep2* detection

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#1 jonbra

jonbra

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Posted 13 June 2017 - 11:54 AM

Sorry for this silly question, but I just wanted to get this right. I am fairly new to miRNA research and I've been using several miRNA-detection tools lately, e.g. mirDeep2* and miR-PREFeR. The problem I have is that all tools seem to generate a whole lot of false positives, even though they follow certain criteria for miRNA detection. For my organism, which is an algae, miR-PREFeR reported more than 2000 candidates, but most of them look like this:

>miRNA-precursor_200 NODE_129_length_55666_cov_574.076:41848-41919 +   
>> Read mappings for sample: thallus 
5'->3'  
UCGAUGUUCGAUGUUCGGUUUCUUGAUGUUUGGUUUACUCCGAAUAUCGAGAAACGAACAUCGAACAUCGU total_mapped_reads=3440 
.((((((((((((((((.(((((((((((((((......))))))))))))))))))))))))))))))). 
....TGTTCGATGTTCGGTTTCTTGATGTT......................................... depth=1, length=26   
.....GTTCGATGTTCGGTTTCT................................................ depth=1, length=18   
.....GTTCGATGTTCGGTTTCTTG.............................................. depth=4, length=20   
.....GTTCGATGTTCGGTTTCTTGA............................................. depth=301, length=21   
.....GTTCGATGTTCGGTTTCTTGAT............................................ depth=10, length=22   
......TTCGATGTTCGGTTTCTT............................................... depth=9, length=18   
......TTCGATGTTCGGTTTCTTG.............................................. depth=52, length=19   
mmmmmmTTCGATGTTCGGTTTCTTGAmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmm depth=1775, length=20 [mature]  
......TTCGATGTTCGGTTTCTTGAT............................................ depth=16, length=21   
......TTCGATGTTCGGTTTCTTGATG........................................... depth=1, length=22   
.......TCGATGTTCGGTTTCTTG.............................................. depth=8, length=18   
.......TCGATGTTCGGTTTCTTGA............................................. depth=177, length=19   
.......TCGATGTTCGGTTTCTTGAT............................................ depth=2, length=20   
........CGATGTTCGGTTTCTTGA............................................. depth=952, length=18   
........CGATGTTCGGTTTCTTGAT............................................ depth=11, length=19   
........CGATGTTCGGTTTCTTGATG........................................... depth=1, length=20   
........CGATGTTCGGTTTCTTGATGTTTGG...................................... depth=1, length=25   
.........GATGTTCGGTTTCTTGAT............................................ depth=86, length=18   
.........GATGTTCGGTTTCTTGATG........................................... depth=2, length=19   
.........GATGTTCGGTTTCTTGATGTTTGGT..................................... depth=2, length=25   
.........GATGTTCGGTTTCTTGATGTTTGGTT.................................... depth=3, length=26   
....................TCTTGATGTTTGGTTTACTCCGA............................ depth=2, length=23   
...........................GTTTGGTTTACTCCGAATA......................... depth=1, length=19   
..............................TGGTTTACTCCGAATATCGAG.................... depth=1, length=21   
..............................TGGTTTACTCCGAATATCGAGA................... depth=3, length=22   
..............................TGGTTTACTCCGAATATCGAGAA.................. depth=1, length=23   
......................................TCCGAATATCGAGAAACG............... depth=1, length=18   
.......................................CCGAATATCGAGAAACGAACATC......... depth=1, length=23   
........................................CGAATATCGAGAAACGAACATC......... depth=1, length=22   
..............................................TCGAGAAACGAACATCGAAC..... depth=1, length=20   
...............................................CGAGAAACGAACATCGAAC..... depth=1, length=19   
...............................................CGAGAAACGAACATCGAACA.... depth=1, length=20   
ssssssssssssssssssssssssssssssssssssssssssssssssGAGAAACGAACATCGAACAssss depth=1, length=19 [star]  
 ...................................................AAACGAACATCGAACATCG. depth=1, length=19   
....................................................AACGAACATCGAACATCG. depth=9, length=18  

However, I found a few looking like this:

>miRNA-precursor_1430 NODE_4145_length_8143_cov_145.377:505-686 - 
>> Read mappings for sample: thallus 
5'->3' ACCGCGAGACUUUGACUUGAAACGGAGGAUUUCUCGAGAUACAAUGACUUCAGUCGUAAAUCGAGGUAUUUUAACGGUUUUCGGGUGUGAUUUUUCACCGAUAUCGUAGGGAAACACCUCGAUUUACGACUGAAGUCAUUGUAUCUCGAGAAAUCCUCCAUUUCAAGUCAAACUCUCGCGG total_mapped_reads=34 
.((((((((.((((((((((((.((((((((((((((((((((((((((((((((((((((((((((.((((.(((((..((((.((........)))))).)))))...)))).)))))))))))))))))))))))))))))))))))))))))))).)))))))))))).)))))))) 
sssssssssssssssssssssssssAGGATTTCTCGAGATACAATGsssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssssss depth=2, length=21 [star] 
..............................................................................TTTCGGGTGTGATTTTTCAC................................................................................... depth=1, length=20 
..........................................................................................................................................TTGTATCTCGAGAAATCCT........................ depth=1, length=19 
..........................................................................................................................................TTGTATCTCGAGAAATCCTC....................... depth=1, length=20 
mmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmTTGTATCTCGAGAAATCCTCCmmmmmmmmmmmmmmmmmmmmmm depth=29, length=21 [mature]

And wouldn't this be a bona fide miRNA? The mature:star forms 2-nt 3' overhangs, and I don't think mismatches between mature and star is a requirement?

 

Thanks for your opinions!




					
					

Edited by jonbra, 13 June 2017 - 11:55 AM.






Also tagged with one or more of these keywords: mir-prefer, mirdeep2*, detection

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