Bisulfite Treatment – Oly Reciprocal Transplant DNA & C.gigas Lotterhos DNA for BS-seq

kubu4 — Fri, 18 Dec 2015 23:23:46 +0000

After confirming that the DNA available for this project looked good, I performed bisulfite treatment on the following gDNA samples:

Sample names breakdown like this:

1NF#

1 = Fidalgo Bay outplants

NF = Fidalgo Bay broodstock origination

# = Sample number

2NF#

Same as above, but:

2 = Oyster Bay outplants

NF2_# (Oysters grown in Oyster Bay; DNA provided by Katherine Silliman)

NF2 = Fidalgo Bay broodstock origination, family #2

# = Sample number

M2/M3 = C.gigas from Katie Lotterhos

Used the EZ DNA Methylation-Gold Kit (ZymoResearch) according to the manufacturer’s protocol with the following changes/notes:

Used 100ng DNA (per Illumina recs; Zymo recommends at least 200ng for “optimal results”).
Thermal cycling was performed in 0.5mL thin-wall tubes in a PTC-200 (MJ Research) using a heated lid
Centrifugations were performed at 13,000g
Desulphonation incubation for 20mins.

DNA quantity calculations are here (Google Sheet): 20151218_oly_bisulfite_calcs

Samples were stored @ -20C. Will check samples via Bioanalyzer before proceeding to library construction.

Sam's Notebook » EZ DNA Methylation-Gold Kit