[BioC] technical vs biological replication in real-time

[r.athanasiadou]

Hi everyone,
My question is of statistical nature.
I am trying to finalize the analysis of real-time PCR experiments.
I have 4 technical replicates (pipeting errors) and three biological
replicates (repeat treatment).

I use gapdh as an external reference gene.

1) Can I average all replication regardless of technical/biological?
2) Similarily can I use the standard error from the combined standard
deviations and what would the population size be in that case? (would it be
4 technical x 3 biological?)

 If I can not do the above how does someone proceed with these two different
types of replication?


I can imagine that the technical replication reflects only on the threshold
cycles and the biological replication at the actual fold
difference[2^-(threshold cycle)] but how can I combine the two differently
calculated standard errors?



I apologize that the e-mail is not directly linking to some BioC package but
there are no packages for real-time and I had nowhere else to turn!

Thanks
Niki




Swann Building
The King's Buildings
University of Edinburgh
Edinburgh, EH9 3JR
Scotland
UK

tel:(0044)0131-6507072
fax:(0044)0131-6505379
R.Athanasiadou at sms.ed.ac.uk



-----Original Message-----
From: bioconductor-bounces at stat.math.ethz.ch
[mailto:bioconductor-bounces at stat.math.ethz.ch] On Behalf Of Pan Du
Sent: 06 July 2007 15:46
To: bioconductor at stat.math.ethz.ch
Cc: Simon Lin
Subject: Re: [BioC] Manufacturer ids in lumiHumanV1

Hi Benjamin

There are potential problems of directly using Illumina Target identifier or
probe identifier because of its imperfect design. For example, there are
duplicated Target IDs in the same chip. The same probe can have different
Target IDs. The nuID is designed to solve these problems. It can be directly
convert to the probe sequence and get the latest annotation easily. Also,
the annotation packages like lumiHumanV1 provides the mapping between
TargetID to nuID and probe Id to nuID.

Please check the vignette of lumi package and the paper published in Biology
Direct 2007, 2:16:
nuID: a universal naming scheme of oligonucleotides for Illumina,
Affymetrix, and other microarrays

Tell me if you have any questions.

Pan



On 7/6/07 5:00 AM, "bioconductor-request at stat.math.ethz.ch"
<bioconductor-request at stat.math.ethz.ch> wrote:

> ------------------------------
> 
> Message: 2
> Date: Thu, 5 Jul 2007 13:00:14 +0200
> From: "Benjamin Otto" <b.otto at uke.uni-hamburg.de>
> Subject: [BioC] Manufacturer ids in lumiHumanV1
> To: "BioClist" <bioconductor at stat.math.ethz.ch>
> Message-ID: <000901c7bef3$ac604870$9f05a20a at matrix.com>
> Content-Type: text/plain; charset="us-ascii"
> 
> Dear bioconductors,
> 
> I just installed the lumiHumanV1 package via the biocLite() procedure
> yesterday. The identifiers of the single probes look very strange to me.
> Beside that they don't appear  in the annotation file provided by Illumina
> the name composition itself made me, let me say somehow suspicious! Here
> comes a quick extract. Did someone observe that before?

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