Question

A Math Problem In Hiseq ??

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11.5 years ago

CrazyB ▴ 280

Not sure if this is the appropriate forum to post this question. If any moderator deems this question inappropriate, please help take it down.

People told me they could do 3 "6-plex" exome sequencing in one Hi-seq sequencing lane, with read-depth of 100x on average. If we assume

1 total exons = 50 MB
6-plex = 300 MB,
3 "6-plex" = 3*300 = 900 MB
100x on average = 900*100 = 90000 MB = 90 GB

Hi-seq sequencing lane has the capacity = 30 GB. So how could they get 3 "6-plex" in one sequencing lane ?

What's wrong with my math here? Any help ? Thanks a lot

ngs hiseq • 2.5k views

ADD COMMENT • link updated 11.5 years ago by matted 7.8k • written 11.5 years ago by CrazyB ▴ 280

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This might help you demistify the maths behind samples and coverage: http://support.illumina.com/downloads/sequencing_coverage_calculator.ilmn

ADD REPLY • link 11.5 years ago by User 59 13k

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Based on the EXCEL spreadsheet, it's not possible to do both 6-plex AND pooling 3 exome seq in one lane, while either-or is feasible. Thanks a lot for the calculator.

ADD REPLY • link 11.5 years ago by CrazyB ▴ 280

score 1 · Answer 1 · 2013-07-09

The math seems fine. Maybe they were using slightly different assumptions? If you say the exome is 30 MB (the size of some selection kits) and the HiSeq lane is getting 250M reads, 120+120bp paired-end, it's fine (you need 54 GB, you get 60 GB off the sequencer). Those are pretty aggressive numbers for a Hiseq, but not unheard of.