The pulverized tissue was placed into a 1.5ml tube containing lysis buVer (10mM Tris–HCl, pH 8.0,100mM EDTA, 0.5% SDS). These samples were thoroughly mixed and incubated overnight at 37 °C. Samples were incubated for 3 h at 37 °C with proteinase K (100 g/ml) and RNase A (20 g/ml), and subsequently extracted with an equal volume of phenol:chloroform:isoamyl alcohol. Total genomic DNA was Wnally precipitated overnight with addition of 1/20 volume 5M NaCl and 2 volume 99% ethanol at ¡20 °C.