Comparison of whole-genome bisulfite sequencing library preparation strategies identifies sources of biases affecting DNA methylation data

Table 2 Library preparation parameters of WGBS strategies compared in this study

Method	Strategy	BS conversion	PCR cycles	Polymerase	Libraries/Studies/Labs
Alkaline	Pre-bisulfite	Alkaline	15–18	Pfu Turbo Cx	18/2/1
Heat	Pre-bisulfite	Heat	10–16	Pfu Turbo Cx	49/4/3^a
KAPA	Pre-bisulfite	Heat and alkaline	6–15	KAPA Uracil+	47/4/2
Am-BS	Pre-bisulfite	Am-BS	9	JumpStart	14/2/1
PBAT	Post-bisulfite	Heat	None^b	–	51/3/2
ampPBAT	Post-bisulfite	Heat and alkaline^c	5–12^b	KAPA HiFi	23/4/2
EpiGnome	Post-bisulfite	Heat	9–10^b	FailSafe	14/2/2

^aOne of the ‘Heat’ studies is generated with Hot Start Taq and is only used as part of Set 2 for Fig. 5 and Additional file 2: Figure S6 and S7
^bInclude one, three or five steps of strand synthesis and pre-PCR enrichment (see Additional file 1)
^cThe alkaline procedure has modifications from the manufacturer’s protocol (see Additional file 1)

ISSN: 1474-760X