Quantitation of next generation sequencing library preparation protocol efficiencies using droplet digital PCR assays - a systematic comparison of DNA library preparation kits for Illumina sequencing

Table 2 Description of the type and number of steps for each DNA library kit tested

	End repair	Bead cleaning	A-tailing	Bead cleaning	Adaptor ligation	Bead cleaning	PCR & bead cleaning	Number of steps after shearing
NEBNext	x	x	x	x	x	x	x	8
NEBNext Ultra	2 in 1 ^a				x	x	x	5
SureSelect	x	x	x	x	x	x	x	8
Truseq Nano	x	x	x		x	x	x	7
Truseq DNA PCR-free	x	xx ^b	x		x	x		6
Accel-NGS 1S ^c	Adaptase		1st extension	x	2nd extension	x	x	7
Accel-NGS 2S ^c	4 different steps + 4 bead cleaning						x	10
KAPA Hyper^d		2 in 1 ^a			x	x	(x)	3 (or 5)
KAPA HyperPlus^d,e			x		x	x	(x)	3 (or 5)

^aBoth End-repair and A-tailing enzymes are combined in a single reaction mix
^bIllumina recommends performing an upper and lower bead clean-up selection after the end repair step
^cSwift Biosciences Accel protocols follow different chemical steps than the classical end-repair, A-tailing, adaptor ligation and PCR
^dKAPA Hyper and KAPA HyperPlus protocol don’t always require a PCR amplification step
^eKAPA HyperPlus protocol starts with non-sheared DNA. The 1st step of the protocol corresponds to the enzymatic shearing of the DNA sample (fragmentase). This fragmentase step leaves blunt-ended DNA fragments which don’t require End-repair and can go straight to A-tailing without any bead clean-up

ISSN: 1471-2164