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ChIP-seq multiplexing - Anyone done it with human samples? (Apr/13/2010 )

Hi everyone :)

Has anyone tried multi-plexing human samples for ChIP-seq? If you have, I have a couple of questions :P

Clare

-Clare-

Hi, I heard that doing multiplex chip-seq on a upgraded solexa can be done for human samples. Since they have had excellent results with sequencing runs generating ~10 million reads. Nowadays a decent center gets about ~30 million reads. So on the safe side 2 human chip-seqs / lane can be done easily. A bit more risk: a good run should take 3 or even 4 chip-seq samples. Be aware that getting the proportions right (same amount of input DNA / sample) is always a tricky business. It depends on the size of your project. If you have only money for a limited amount of lanes and nothing to spare on doing an extra lane when the datayield is low for a run keep it conservative...
Chip-bisulfite sequencing multiplexing is definetely still a no go.

best
E

-et2b-

Thanks for the reply :)

We're thinking about multiplexing as we have at least 14 samples that we want to Chip-seq. We ran a pilot 2 lanes and got 7 and 9 million reads that aligned. With this data we randomnly discarded 60% of the reads (to mimic multiplexing) and then looked at the results... It seems we do lose a lot of peaks with ~ half the number of reads :D BUT if we had access to one of the newer sequencers we may be ok...

hmm...

I'll keep the forum posted with what happens :P

Have a great weekend!

Clare

et2b on Apr 16 2010, 10:04 AM said:

Hi, I heard that doing multiplex chip-seq on a upgraded solexa can be done for human samples. Since they have had excellent results with sequencing runs generating ~10 million reads. Nowadays a decent center gets about ~30 million reads. So on the safe side 2 human chip-seqs / lane can be done easily. A bit more risk: a good run should take 3 or even 4 chip-seq samples. Be aware that getting the proportions right (same amount of input DNA / sample) is always a tricky business. It depends on the size of your project. If you have only money for a limited amount of lanes and nothing to spare on doing an extra lane when the datayield is low for a run keep it conservative...
Chip-bisulfite sequencing multiplexing is definetely still a no go.

best
E

-Clare-

Hi Clare,

please be aware there is a seperate forum on next gen. sequencing. Its ideal to get help of people currently doing experiments and more importantly people working on the same sequencing platform.

best

-et2b-

Hi,

Are you talking about this one?

http://seqanswers.com/

I'm a one forum kinda gal but I'll have a look :P Thanks!

Clare

et2b on Apr 20 2010, 02:07 PM said:

Hi Clare,

please be aware there is a seperate forum on next gen. sequencing. Its ideal to get help of people currently doing experiments and more importantly people working on the same sequencing platform.

best

-Clare-