S.N. Components/Steps Method 1 Method 2 Method 3 Method 4
1 CTAB 2% 2% 2.5% 2.5%
2 Sonication 3 cycles at high amplitute No 3 cycles at high amplitute No
3 FREEZE-THAW 3 cycles at 70°C and -20°C 3 cycles at 70°C and Liquid Nitrogen 3 cycles at 70°C and Liquid Nitrogen No
4
FREEZE-BOIL
No
No
No
3 cycles at 95°C for 5 minutes, -80°C for 10 minutes
5 DNase-Free-RNase No No 10µl/ml 10µl/ml
6 Chloroform: isoamyl alcohol (24:1) ~800 µl ~800 µl ~800 µl ~800 µl
7 Phenol: Chloroform: isoamyl alcohol (25:24:1) ~600 µl ~600 µl ~600 µl ~600 µl
8 Isopropanol ~400 µl ~400 µl ~400 µl ~400 µl
9 Ice-cold Ethanol 70% 1ml, twice 1ml, twice 1ml, twice 1ml, twice
10 Nucleus Free Water 30 µl 30 µl 30 µl 30 µl
11 18ssu rRNA PCR No No Yes Yes