Read count variation

[Aihui Wang]

Hi Joe,

We performed LCM FFPE followed by PreSPRI pooling for library preparation. When we look at the expression matrix, some samples have hundreds of readcounts while others have 1 or 2, or zero, even for housekeeping genes.  I wonder if it is normal to see read counts varying a lot.

Thanks,

Aihui

[Joe Foley]

Generally that sounds normal. There's always a little bit of variation in yield from one library to the next, and when you work directly on FFPE LCM tissue that variation can be extreme, with some samples occasionally failing altogether. When you do the pre-SPRI pooling, you can't rebalance the libraries to compensate for that variation, so you're just relying on luck and hopefully enough replicates that you can afford to lose a few.

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