Hi Peter,
For more information on what Mike explains, I recommend this paper:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2773710/

Also, be careful about distinguishing whether your FDR<=1% is at the unique 
peptide level or the peptide spectrum match (PSM) level.  The same 
phenomenon occurs between PSM FDR and unique peptide FDR as between unique 
peptide FDR and protein FDR.  Namely, true discoveries are more likely to 
be repeated, and therefore 1% FDR at the PSM level will grow above 1% for 
unique peptides and again further for proteins, as Mike noted.

Your choice to include only proteins with >1 hit does improve this growth 
in FDR, by applying a secondary filter, but it is by no means foolproof 
with modern mass spectrometers and multi-run data sets, depending on the 
number of spectra you collected versus the number of proteins considered in 
your search.  I have seen million spectrum data sets where FDR was 
seriously underestimated against a human FASTA with proteins for ~23,000 
genes.

A good way to gain some insight on your true FDR is to have decoy proteins 
in your FASTA that are never exposed to any of your search criteria.  This 
allows you to double-check FDR estimates from the statistical tools you 
use.  Once you gain familiarity with your tools, this may be come less 
necessary, but I have seen statistical tools get these numbers wrong, for 
exactly these reasons.

Good luck with your FDR statistics.

--Brendan

On Tuesday, November 26, 2013 8:52:25 PM UTC-8, zion wrote:
>
> Dear All:
>
> If there are some confident peptide ids at FDR<=1% at the peptide level, 
> when inferring proteins by confident peptides and at least one unique 
> peptide, CAN i say that at the protein level, its FDR<=1%?
> Many thanks,
>
> -Peter
>

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