Multiplex more than 12 samples?

[cybeline]

Hello,

I was wondering if it was possible to multiplex more than 12 samples per lane?
It seems most people only multiplex 12 and if it is possible to do more what are the cons?

Thank you,

[DMO]

You can index many more than 12. We have designed primers for 96 indexes and have been using them for the last 6 months or so with very good results (nice even representation across all 96). For many projects indexing 96 samples in a lane may not be suitable because you may need more coverage than that. But, with the ever increasing popularity of targeted capture of small regions and as the # of reads/lane doubles or triples over the next year (Illumina is getting 1.14TB/2x150bp run on a HiSeq2000(2flowcells)) , indexing many more than 96 will soon become very popular.

[cybeline]

Thank you very much for your answer.

Could you please expand a little on the coverage issue with multiplexing 96 samples or more than 12 in general? What type of results would you expect per sample? how many sequences read are you able to get?

Thanks again,

[DMO]

Well, you still get the same average number of reads per lane, just divvied up among the respective samples in the lane. So you just need to figure out how many reads you need for each sample and pool it in at whatever % of a lane you need for that sample.

[cybeline]

Makes sense, thank you again!