DNA library 2 from pubic hair 1 of individual HP1-S003 (Hair1.0 panel): Sample JK-P1-S003
Pubic hair was trimmed to <3cm and root removed. For decontamination, hair was submerged in 0.5% sodium hypochlorite followed by water baths. DNA was extracted using the Qiagen DNeasy Blood and Tissue Kit following a Qiagen user-developed protocol for hair and eluted in 40 µL buffer EBT. Illumina library was created by concentrating 20 µL of the extraction to 11 µL with a SPRI bead mixture. We then prepared the library as described in Kapp et al. (2021) with some modifications. Next, library was amplified and double-indexed using primers described in Kircher et al. (2012). We prepared 50 µL reactions containing 20 µL library, 25 µL Amplitaq Gold 360 Master Mix (Applied Biosystems), 2.5 µL unique 20 µM i5 indexing primer, and 2.5 µL unique 20 µM i7 indexing primer. Then, we amplified the library with the following cycling conditions: 95°C for 10 min, followed by 10 or 13 cycles of 95°C for 30 s, 60°C for 30 s, and 72°C for 60 s, and a final extension of 72°C for 7 min. Finally, we purified the hair library using a SPRI ratio of 1.2X and sequenced at UCSF on one lane of NovaSeq using the S4 2x150 kit.