Pooled the libraries into a single sample for sequencing on Illumina HiSeq2500 by Genewiz.
Here’s the list of samples that were pooled:
| SAMPLE NAME | LIBRARY NAMES | INDEX 1 (i7) | LENGTH (bp) | INDEX 2 (i5) | LENGTH |
| SJW_Oly_2bRAD | Oly RAD 02 | CGTGAT | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 03 | ACATCG | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 04 | GCCTAA | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 06 | TGGTCA | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 07 | CACTGT | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 08 | ATTGGC | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 14 | GATCTG | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 17 | TCAAGT | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 23 | CTGATC | 6 | ATGCAT | 6 |
| SJW_Oly_2bRAD | Oly RAD 30 | AAGCTA | 6 | ATGCAT | 6 |
Combined 40ng of all samples, except Oly RAD 30. Used only 20ng of Oly RAD 30 because it was the only sample that produced a single peak in qPCR melt curve analysis (i.e. no primer dimer). As such, it’s a rough assumption that the qPCR quantitation of all the other samples is twice as high as they should be due to the contribution of primer dimer amplification.
Calculations for pooling can be seen here (Google Sheet): 20151117_RAD_qPCR_data
The full list of samples (and the individual samples/libraries/indexes) submitted to Genewiz for this project by Katherine Silliman & me can be seen here (Google Sheet): White_BS1511196_R2_barcodes