Four representative S. saccata specimens were selected and
used for further molecular analyses. Lichen thalli with apothecial discs were mainly used
for DNA extraction. Samples were ground and extracted with DNeasy plant mini kit (Qiagen,
Valencia, CA, USA) according to the manufacturer’s instructions. PCR amplifications were
conducted using Amplitaq DNA polymerase (ThermoFisher, Watham, MA, USA). The following
primers were used for PCR amplifications: mtSSU1 and mtSSU3R for mtSSU [29 ]; NS17UCB, NS20UCB for nuSSU [30 ]; LIC24R [31 ] and LR7 [32 ] for nuLSU; fRPB2-7cf
and fRPB2-11aR for RPB2 [33 (link)]. PCR conditions
for nuSSU are as described in a previous study [34 (link)]. The following program was used for amplification of nuLSU: initial
denaturation for 4 min at 94 °C, followed by 30 cycles of 94 °C for 40 s, 52 °C for 40 s,
72 °C for 50 s, and then a final extension step at 72 °C for 8 min. Amplified DNA was
concentrated and purified using a PCR quick-spin PCR Product Purification Kit (INTRON
Biotechnology, Inc. Sungnam City, Korea) for sequencing analysis.
used for further molecular analyses. Lichen thalli with apothecial discs were mainly used
for DNA extraction. Samples were ground and extracted with DNeasy plant mini kit (Qiagen,
Valencia, CA, USA) according to the manufacturer’s instructions. PCR amplifications were
conducted using Amplitaq DNA polymerase (ThermoFisher, Watham, MA, USA). The following
primers were used for PCR amplifications: mtSSU1 and mtSSU3R for mtSSU [29 ]; NS17UCB, NS20UCB for nuSSU [30 ]; LIC24R [31 ] and LR7 [32 ] for nuLSU; fRPB2-7cf
and fRPB2-11aR for RPB2 [33 (link)]. PCR conditions
for nuSSU are as described in a previous study [34 (link)]. The following program was used for amplification of nuLSU: initial
denaturation for 4 min at 94 °C, followed by 30 cycles of 94 °C for 40 s, 52 °C for 40 s,
72 °C for 50 s, and then a final extension step at 72 °C for 8 min. Amplified DNA was
concentrated and purified using a PCR quick-spin PCR Product Purification Kit (INTRON
Biotechnology, Inc. Sungnam City, Korea) for sequencing analysis.
