Hi all,
To reach reasonable DNA concentrations to put my pooled libraries on MiSeq I need to concentrate my samples i.e. my pooled libraries.
I have a huge range of DNA concentrations over the ~100 samples I am going to pool.
Through equimolar combination and dilution I will end up with 200ul of sample with a concentration at least 10x too low (around 0.2nM)!
Concentrating through SpeedVac is no option since my samples are in buffer and not in H2O.
I was thus planning to concentrate the pool with one step of Ampure beads and eluting them in 10x less volume. So I would elute 40ul from 200ul sample + 200ul bead to ideally rech a concentration of ~2nM.
Does anybody have experience with this? Would that work? How big of a loss do I have to expect?
Thanks a lot for your help!
jdp
To reach reasonable DNA concentrations to put my pooled libraries on MiSeq I need to concentrate my samples i.e. my pooled libraries.
I have a huge range of DNA concentrations over the ~100 samples I am going to pool.
Through equimolar combination and dilution I will end up with 200ul of sample with a concentration at least 10x too low (around 0.2nM)!
Concentrating through SpeedVac is no option since my samples are in buffer and not in H2O.
I was thus planning to concentrate the pool with one step of Ampure beads and eluting them in 10x less volume. So I would elute 40ul from 200ul sample + 200ul bead to ideally rech a concentration of ~2nM.
Does anybody have experience with this? Would that work? How big of a loss do I have to expect?
Thanks a lot for your help!
jdp
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