DNA Methylation Test - Gigas site gDNA (BB & DH) from 20090515

Used BB & DH samples #11-17 for procedure. Followed Epigentek’s protocol. My calcs for dilutions/solutions needed are here. All solutions were made fresh before using, except for Diluted GU1 which was made at the beginning of the procedure and stored on ice in a 50mL conical wrapped in aluminum foil. Used 100ng total (50ng/uL) of each sample gDNA. No standards for a standard curve based on speaking with Mac.

A01 BB11 A02 DH11
B01 BB12 B02 DH12
C01 BB13 C02 DH13
D01 BB14 D02 DH14
E01 BB15 E02 DH15
F01 BB16 F02 DH16
G01 BB17 G02 DH17
H01 Pos. Control H02 Blank

Results: Above is the graph of the results. Although it’s only a small difference between the two sites, it is statistically significant. [The calcs for this graph can be found here (Excel file)(https://eagle.fish.washington.edu/Arabidopsis/Notebook%20Workup%20Files/20090519%20Gigas%20DNA%20methylation%20workup%20SJW.xls). It should be noted that this graph was generated using estimated values from the standard curve provided in the manufacturer’s protocol. This was done because 1) I did not run standards to generate my own curve and 2) calculating the “% methylation” not using the formula that utilizes the standard curve was giving ridiculously high values (e.g. 350%).

Here is the raw data generated by the plate reader for a [1s read (Excel file)(https://eagle.fish.washington.edu/Arabidopsis/Notebook%20Workup%20Files/20090519%20gDNA%20Methylation%201s%20SJW.xls) and a 0.1s (Excel file) read. Both reads have nearly identical values.