Shelly’s Notebook: Tues. May 14, 2019 Salmon-sea lice salinity x temp methylomes

Extracted gDNA from salmon skin samples

These samples are from Christian and are from animals that were infected with sea lice for the duration of the sea lice life cycle in two different temperatures (8C and 16C) and two different salinities (26psu and 32 psu). Sample info here

Kaitlyn helped with the extraction.

Followed the EZNA Mollusc DNA kit protocol

  • weighed out 40-50mg tissue and added to microfuge tube
    • remaining ethanol-fixed tissue is in a box on the bottom shelf of the 4C in 213
  • froze samples in liquid N2 and used disposable microtube pestle to pulverize sample
    • this was a little challenging as the pestle didn’t really seems to break up the tissue, just squish it. Also used pipette tip to smash tissue further.
    • samples sat on ice after being pulverized until we got through all the samples
  • incubated samlpes in 305uL ML1 buffer and 25uL Proteinase K solution for 1 hour at 60C
    • not all the sample was solubilized but a lot of it was
  • did the optional column equilibration step
  • eluted in 50uL elution buffer for 5 min at RT before spinning for 2 min at 10,000xg

DNA concentration

  • Qubit dsDNA BR kit for 20 samples + 2 standards
    • Ran Standard initially read 99.7ng/uL
      • read it again after every 5 samples and it consistently read 102ng/uL
    • sample concentrations are here: 20190408_SalmonSamples
      • lowest yield was 61ng/uL * 50uL = 3050ug so we have plenty of DNA
  • DNA are in a box in the -20C in 213 in the bottom drawer

Next steps

  • Digest 1ug of each sample with MspI and Taq-a-1 (o/n)
  • Zymo pico methyl library prep kit on salmon samples and sea lice samples

from shellytrigg http://bit.ly/2Hk9e7d
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