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Old 11-24-2015, 01:36 PM   #1
AaronReedy
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Location: Virginia, USA

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Default Limits to pooling large number of samples?

Iím working on a ddRADseq project. We are planning to sequence 2,800 individual lizards from a closed island population. Our plan is to use 48 separate inline barcodes and 59 pcr indices to identify individual samples. We digested with EcoRI and SphI and after size selection expect 7-8,000 fragments per individual. The current plan is to combine samples from all 2,800 individuals into a single pool and sequence all individuals in each HiSeq lane. We will then run as many lanes of sequencing as needed to attain desired coverage (~20x)?

Does pooling this number of individuals seem reasonable to people with experience on ddRAD projects with large numbers of samples?
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Old 11-24-2015, 02:21 PM   #2
nucacidhunter
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If 59 indices have good distance from each other for correct demultiplexing I do not see any issues. You just will have a lot of work merging reads from each sample into a single file after demultiplexing with indices and inline barcodes.
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Old 03-17-2016, 07:17 AM   #3
InDel
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There is no limit for pooling samples.
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Old 03-21-2016, 01:59 PM   #4
AaronReedy
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Default Thanks.

Thanks for the answers. We are moving ahead with the project as planned.
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