[Morgane_AUS]

We have the Qsep (similar to bioanalyser) but something went wrong with the cartridge so I had to use a gel. I had to overexpose it in order to see the smear (there is only 1 uL of ladder..).

I will include a no template library prep next time, I couldn't here because of the number of samples I have and kit size.

may I ask why you would do the 0.9x bead twice ?

Thanks for your insights !

[nucacidhunter]

One time wash less likely will get rid of that band. If a small residue of that band (prsumably primer- or adapter-dimer) is left in youer sample, they will form clusters more efficiently and you may get very small portion of reads from your target fragments.

[Morgane_AUS]

I don't have much DNA in there so I'm worried I will loose so much by doing it twice... I guess there is no win win situation here ! Finger crossed !

[nucacidhunter]

I do not know how many PCR cycles you have done. But you can do 2-3 cycles after 1st clean up.

[cmbetts]

I was about to suggest the same thing. Post-PCR, you always have the option of hitting it with a few more cycles if you end up with low yield. At this point, all of the fragments have been amplified enough that you don't really need to worry about drop-out anymore, you just run the risk of further distorting their relative abundance.