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Topics - wedmands

1
metaXCMS / P-Value of zero in common features?
I am observing a p-value of zero in my metaXCMS common features? It is obviously incorrect to report this p-value..Is there a certain number of decimal places which metaXCMS reports? Are these values indeed significant or not? Has anyone else seen this?

Thanks in advance.  :)  :)
2
metaXCMS / When to normalise urine using metaXCMS?
Hi,

I am using metaXCMS for human urine samples where there are 3 subjects and 2 conditions (fed and unfed) with 3 replicate injections and I need to normalise appropriately to account for sample dilution between subjects. It would be good to normalise across all of the samples together to the median fold change and this would usually be following peakpicking. However if I am treating each subject individually I cannot easily do this, perhaps there is a way as I don't want any false negatives/positives. I don't know how I could do this when I am generating a diffreport seperately for each individual? I read through the nature protocols paper and could not find any suggestions on fitting in a normalisation step. I have already used metaXCMS and it is working very well and some interesting features have arisen based on monoisotopic mass but I want to ensure my interpretation is accurate.

I am considering in future to normalise to osmolality and correcting each sample prior to data acquistion in future however for now does anyone have any suggestions of how I can normalise these urine samples so that I don't misintepret the second order analyses with metaXCMS and fit it into the work scheme?

Also does anyone have any suggestions on how I can search my own database or the soon to be released FooDB when generating the XCMS diff report instead of/ in addition to Metlin?

Thanks very much.  :)
3
CAMERA / GroupCorr algorithm error
I am trying to annotate adducts using CAMERA from urinary data acquired using an UPLC-Agilent 'ifunnel' QTOF 6550 in centroid mode (converted to mzXML using msconvert)and I am having difficulty grouping correlated peaks with the groupCorr algorithm. I would really appreciate any help as I am a little bit stuck. Sorry for all the information but I thought it best to provide as much as possible. :)

I recieve this error message and believe it might be due to my memory usage: -

>xsIC <- groupCorr(xsI, cor_eic_th = 0.75)
Start grouping after correlation.
Generating EIC's ..
Error: cannot allocate vector of size 3.6 Gb

where xsl is an xsAnnotate object of: -

With 1028 groups (pseudospectra)
With 18 samples and 62755 peaks
Polarity mode is set to: 
Using automatic sample selection
Annotated isotopes: 8739
Memory usage: 177 MB

Prior to this
> xsF <- groupFWHM(xsa, perfwhm = 0.6)
Start grouping after retention time.
Created 1028 pseudospectra.
and....
xsI <- findIsotopes(xsF, mzabs = 0.01)
Generating peak matrix!
Run isotope peak annotation
Found isotopes: 8739


I am using a virtual machine with the following characteristics and I hoped these would be sufficient

Windows 7 64 bit
8GB of RAM
2 CPUs
1 System disk of 40GB
1 Data disk of 1TB

I wonder if the number of peaks picked using XCMS is rather a lofty figure? I put this down to being very high resolution spectra, (as there were some peak insertion problems using ppm values of 30 and 20) long gradient and EICs from the diffreport appeared satisfactory, perhaps I have gone wrong somewhere with my peak picking, peak grouping and retention time correction prior to using CAMERA.

My XCMS workflow consisted of: -

>peakmatrix <- xcmsSet(method="centWave", peakwidth=c(3,20), ppm=15, snthresh=10);
An "xcmsSet" object with 18 samples
Time range: 3.4-1500.1 seconds (0.1-25 minutes)
Mass range: 100.0022-1049.6504 m/z
Peaks: 836770 (about 46487 per sample)
Peak Groups: 0
Sample classes: A1, A2, B1, B2, J1, J2
Profile settings: method = bin
                  step = 0.1
Memory usage: 72.3 MB

>gs<-group(peakmatrix,bw=10)
gs
Peaks: 836770 (about 46487 per sample)
Peak Groups: 38072
Memory usage: 81.4 MB

Retention  time correction 1st pass
ret<-retcor(gs, p="m", f="s", missing=1, extra=1, span=0.2)
Retention Time Correction Groups: 855
ret
Peaks: 836770 (about 46487 per sample)
Memory usage: 79.7 MB
[attachment=1:2o7a2vxc]Retention_time_correction_1st_pass.jpg[/attachment:2o7a2vxc]

2nd Grouping
gret<-group(ret, bw=5)
gret
Peaks: 836770 (about 46487 per sample)
Peak Groups: 53400
Memory usage: 91.2 MB

Retention time correction 2nd pass
Ret2nd<-retcor(gret, p="m", f="s", missing=1, extra=1, span=0.2)
Retention Time Correction Groups: 1552
Peaks: 836770 (about 46487 per sample)
Memory usage: 79.7 MB
[attachment=0:2o7a2vxc]Retention_time_correction_2nd_pass.jpg[/attachment:2o7a2vxc]

3rd Grouping
gret2nd<-group(ret2nd,bw=3)
Peaks: 836770 (about 46487 per sample)
Peak Groups: 62755
Memory usage: 92.6 MB

Zero filling
fill<-fillPeaks(gret2nd)

Many thanks for your help!

[attachment deleted by admin]
4
XCMS / centWave sort assumption violation
Hi,

I have been having trouble with the centWave peak picker I continually get the following error message at the very end of the process?
Error in checkForRemoteErrors(val) :
  100 nodes produced errors; first error: m/z sort assumption violated ! (scan 611, p 3578, current 322.1408 (I=15.85), last 322.1423)

I have tried peak picking with each individual sub folder in turn to see if one of the folders contains a corrupt file or maybe something else but I still get the same error.

Using the default XCMS not centWave it seems to work fine strangely. I also converted my files using the Databridge programme to netCDF files if this is pertinent information.

This is the command I utilised I set the CPWmin, CPWmax and snthresh beforehand.
>peakmatrix <- xcmsSet(method="centWave", peakwidth=c(CPWmin,CPWmax), ppm=ppm, snthresh=xsnthresh, nSlaves=8)

Apologies for back to back help requests I am new to R and using XCMS and I am not a computational biologist by any means. I wonder if anyone has any ideas?

Will
5
XCMS / Uploading error message?
Hi I was wondering if anyone has come across this error before when uploading your cdf files. The upload commences and completes 50 or so files but always seems to halt on this particular file? I'm puzzled. Thanks for your help. :)

ASP_P_P2_5901:
Traceback:
 1: .C("ProfBinM", x, y, as.integer(length(x)), as.integer(zidx),    as.integer(length(zidx)), as.double(xstart), as.double(xend),    as.integer(num), out = doubleMatrix(num, length(zidx)), NAOK = NAOK,    DUP = FALSE, PACKAGE = "xcms")
 2: profFun(object@env$mz, object@env$intensity, object@scanindex,    bufsize, mass[1], mass[bufsize], TRUE, object@profparam)
 3: .local(object, ...)
 4: findPeaks.matchedFilter(<S4 object of class "xcmsRaw">)
 5: findPeaks.matchedFilter(<S4 object of class "xcmsRaw">)
 6: do.call(method, list(object, ...))
 7: .local(object, ...)
 8: findPeaks(lcraw, ...)
 9: findPeaks(lcraw, ...)
10: xcmsSet()
Error in profFun(object@env$mz, object@env$intensity, object@scanindex,  :
  caught access violation - continue with care
In addition: Warning message:
In xcmsRaw(files, profmethod = profmethod, profparam = profparam,  :
  There are identical scantimes.