Project: Thermal Transplant Molecular

Goal

Eighth trial of WGBS library prep using the Zymo Pico Methyl-seq library prep kit and this protocol. I am re-doing 3 samples that had a PCR bubbles due to over amplification from this post and 2 samples that did not reamplify from this post.

Sample dilution and mix calculations can be found on this google sheet

Protocol

I am using a modified version of this protocol.

For all samples, I used the following modifications:

• Use the combined i5 and i7 index primers (an input of 1uL)
• Reduce the Library Amp Master Mix (2x) to 13uL (was originally 14uL)

Samples and Indices

Sample	Coral ID	Life Stage	i5 Index #	i7 Index #
18-358	P-10-A	Adult	49	49
18-20	P-6-B	Adult	50	50
L-933	P-10-A	Larvae	51	51
L-924	P-9-A	Larvae	52	52
L-1257	P-19-A	Larvae	53	53

Qubit Results

To test DNA quantity: Qubit

I used the DNA broad range assay.

Sample	DNA 1 (ng/ul)	DNA 2 (ng/ul)
Standard 1	180.74
Standard 2	21047.36
49	5.96	5.80
50	11.9	11.7
51	18.1	18.2
52	2.50	2.46
53	20.2	19.7

Tapestation Results

D5000 Tapestation protocol

Full Tapestation Report