What is pooled samples in NGS?

Question

[897598644]

Excuse me:

In the GATK forum, there were following descriptions:

If you are limited to older versions for project continuity, you may opt to use UnifiedGenotyper in the following cases:

• If you are working with pooled samples (also due to the HC's limitation regarding ploidy)

So how could we understand the phase of pooled samples? Was that DNA mixture for multiple samples?

Many thanks in advance!

[andrew.j.skelton73]

Just as a side note, that sounds like an old forum post. Pooling samples is still applicable in the latest GATK best practises, however only in some specific cases.

Answer 1

[apelin20]

You have multiple individuals in one library. Sometimes this can be unavoided, sequencing spores for example of bacteria, or spores of fungi, or populations of viruses infecting one tissue. But sometimes people actually pool DNA, like taking 10 human individuals and pooling their DNA into one sample and sequencing it.

[897598644]

What is the usual purpose if we take 10 human individuals and pooling their DNA into one sample and sequencing it?

[apelin20]

REDUCE COSTS! :) Sequence one sample as opposed to 10. Library prep can be quite expensive.

[andrew.j.skelton73]

and then you destroy any between sample variance - which isn't a good thing most of the time.

[apelin20]

Of course there is a downside. However, an experiment is better than no experiment.

[andrew.j.skelton73]

Pooling human samples is only a good thing if they're genetically identical samples, i.e. I have two technical replicates of the same sample. I've not seen documentation to suggest how GATK would interpret pooled samples in the latest version, i.e. how it would deal with more than two genotypes, if it just calls the sample multi allelic, or something different. In differential expression tests, pooling everything destroys between sample variance, which means statistically you can't estimate the variance of your population.