Tag Archives: EZ DNA Methylation-Gold Kit

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

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

  • 1NF11
  • 1NF15
  • 1NF16
  • 1NF17
  • 2NF5
  • 2NF6
  • 2NF7
  • 2NF8
  • NF2_6
  • NF2_18
  • M2
  • M3

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

 

Followed the guidelines of the TruSeq DNA Methylation Library Prep Guide (Illumina).

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.

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