Kapa hifi hs

TdT-Tailing and pre-amplification reaction were adopted from TELP with modifications^{43 (link)}. 1 μL 10X TdT buffer, 0.5 μL 1 mM dCTP (NEB, N0447S) was added into 12.5 μL purified DNA/cDNA mix. The samples were incubated at 95 °C for 5 min and quickly chilled on ice for 5 min. 1μL of TdT (NEB, M0315S) was then added and tailing reaction was carried out under 37 °C for 30 min followed by inactivation at 75 °C for 20 min. Anchor Mix (6 μL 5X KAPA Buffer, 0.6 μL 10 mM dNTPs, 0.6 μL 10 μM Anchor-Oligo [Supplementary Table 1] and 0.6 μL KAPA HiFi HS [KAPA, KK2502]) were added and the linear amplification was performed with the following program (Step1: 98 °C × 3 min; Step2: 98 °C × 15 s, 47 °C × 60 s, 68 °C × 2 min, 47 °C × 60 s, 68 °C × 2 min and repeat Step2 for additional 14 times; Step3: 72°C × 10 min and hold at 12 °C).
Preamplification Mix (4 μL 5X KAPA buffer, 0.5 μL 10 mM dNTPs, 2 μL of 10 μM PA-F and PA-R [Supplementary Table 1], 0.5 μL KAPA HiFi HS) were then added and preamplification were performed as the following program (Step1: 98 °C × 3 min; Step2: 98 °C × 20 s, 65 °C × 20 s, 72 °C × 2.5 min and repeat Step2 for additional 9 times; Step3: 72°C × 2 min and hold at 12 °C). Amplified products were purified with SPRI double-size selection (10 μL + 32.5 μL) and were eluted in 34 μL ultrapure H₂O, use 1 μL for quantification.