according to the manufacturer’s instructions. Hybridization
was performed for 20 h at 42 ◦C; the membrane was washed
twice in 2×SSC/0.5% SDS solution at room temperature for 5 min,
then twice in 2×SSC/0.1% SDS solution at 68 ◦C for 15 min, followed
by 2×maleic acid/0.6% Tween 20 washing buffer for 5 min.
The membrane was then soaked in detection buffer with NBT/BCIP
solution and exposed to film for 4–12 h