Low yields with HyperPlus library kit

[jmc887]

Hello everyone,

I am trying out the HyperPlus kit for library prep. The protocol says the expected pre-capture yield should be ≥ 1 ug. However, I got around 500 ng yield (my input was 100 ng). I have performed the protocol precisely, so I would like to know if anyone has more experience with this kit and if you have encountered the same issue?

Thanks in advance,

[luc]

How many PCR cycles did you run? Do the libraries have the expected length?

[jmc887]

I ran 7 cycles in total (as indicated for SeqCap adapters), and yes, the libraries have the expected length. Its only the yield that is the lowest I get, compared to other library prep kits using the same amount of input DNA (SureSelect and Nextera Flex).

[jhalpin]

How did you measure your 100ng input?

[jmc887]

How did you measure your 100ng input?

I measured with Qubit

[luc]

Sorry, I have no clue what could have gone wrong. We do see significantly higher yields with the same Kapa kit using Biooscientific adapters after 7 PCR cycles.
Did you run a two-sided size selection? This will significantly reduce yields.

[jmc887]

Thanks for your answer luc, in fact yes, i did run a double-sided size selection. I will probably try again without this double-sided selection.