Recent studies discovered that there are copy number variations in normal tissue samples across individuals (Iafrate et al. 2004, Sebat et al. 2004). The implication is that we cannot always assume that the reference normal samples used to obtain reference signal distribution all have copy number of 2 at a SNP locus. In addition, researchers often cannot obtain enough (~10) normal reference samples to analyze together with tumor samples. When normal samples are not available or too few, set “Options/Chromosome/% of samples trimmed” to be > 0 to obtain reference signal distribution without using the information of which samples are normal. For example, if 10% is specified, we assume that for any SNP less than 10% of all the samples have abnormal copy number. For a SNP, 5% of samples with extreme signals are trimmed from each end, and the rest values are used to estimate the mean and standard deviation of the signal distribution of normal copy number 2. The large amplifications and homozygous deletions can be found, but the method may infer false copy number changes in normal samples. You can try different trimmed % and compare the results.

If you specify "% of samples  trimmed " as a negative value (e.g. -20), only the normal samples (specified as 2 for "Ploidy(numeric)" in sample information file), instead of all samples in the group, are trimmed by this percent and used for computing signal distribution of copy 2.

Figure below: median-smoothed copy number (window size 11 SNPs, trimmed 1% samples) for chromosome 19 of Affymetrix HapMap trio dataset. A sample (name highlighted in blue) has heterozygous deletion in the 19p13.11 region, which is also one of the most frequently deleted region found by Iafrate et al. 2004.

When a region has copy change in the same direction in most samples, this trimming method will likely miss the region. In such case having at least one normal sample will be useful, so at a first round analysis one can pair this normal with every tumor in array list file and check “Options/Use paired normal as reference” to get raw copy number and assess whether such regions are likely.

[Discussion]