I have recently gone through the generation of a ChIP-seq library using the standard Illumina protocol. According to this, I have done two gel purifications, one immediately following the adapter ligation and one immediately following the final library amplification. The library generation seemed to work, as I was able to see bands in the desired size range, however after the final gel purification I was lucky if I had 1ng of total DNA. This is far below what the core facility likes to have (~50ng).
I would love any stories/solutions that anyone can share regarding how much DNA they typically get back? Do you do two gel purifications? How many PCR cycles in the library amplifcation (the protocol calls for 18 cycles, but I have some colleagues that claim 25 is needed).
I would love any stories/solutions that anyone can share regarding how much DNA they typically get back? Do you do two gel purifications? How many PCR cycles in the library amplifcation (the protocol calls for 18 cycles, but I have some colleagues that claim 25 is needed).
Comment