Hi

We have two experiments run on illumina arrays done by the same company. I want to combine the data. I normally use Limmas functions (read.ilmn, and neqc) to deal with normalisation of the data. However if I combine these two experiments what is the best way of going about doing this. I am aware they need background correcting and quantile normalisation using the negative and positive control probes on the arrays.

edit - platform is the same, but need to compare healthy in one group with diseased in the other group. why it was done like this - well i didnt have any say in it - i cant go into that. the company we use is highly efficient and everything is standardised so batch should not be much of a problem.

Best,

Chris

You can simply combine the datasets before normalization like this:

x1 <- read.ilmn( files for run1 )
x2 <- read.ilmn( files for run2 )
x <- cbind(x1,x2)
y <- neqc(x)

etc. This assumes that the probes for both runs are identical and in the same order.

Dear Chris,

actually the the task of combining 2 or more datasets is always challenging, and also with many parameters or assumptions to take into account !! Firstly, the illumina arrays are on the same platform, or different ? Secondly, the phenotype information or the experimental design is the same ? For istance, the same cell line used or different? Then, it would also be the presence of a possible batch effect, that you will have to take into account. Thus, firstly -in my personal opinion- the safest way is to analyze separately the two datasets, and then:

1) Compare your final DE lists, to identify common genuine DE genes among your "similar comparisons";

2) Identify common pathways or ontologies "enriched" in both datasets;

3) Also perform other exploratory plots of the DE statistics, such as a scatter plot of Log-FCs.

Then, it might be interesting for other reasons (i.e. for gaining theoretically more statistical power) to combine both datasets. But still, there are a lot of issues that you will have to take into account, in order even for the combination to be valid. I hope that helps.

Best,

Efstathios