DNA extraction (BOMB)
Yin-Tse Huang, Tsu-Chun Hung
Abstract
DNA extraction (BOMB)
Steps
Sample Collection
Add 200µL of 1mm beads to 1.5mL eppendorf tube
Add200µL of 0.5mm beads to 1.5mL eppendorf tube
Add870µL Lysis master mix to 1.5mL eppendorf tube
Collect 10-20mg of sample to 1.5mL eppendorf tube
Sample crush
Put 1.5mL eppendorf tube in mixmill for sample crush, at this condition: 30 rpm/s, for 4mins 0h 4m 0s
Centrifugation
Put 1.5mL eppendorf tube in centrifuge for centrifugation, at this condition:17.0x g,25°C
DNA purification
Add 350µL of isopropanol to the 1st well of 96 well plate
Add 100µL of magnetic beads (10 mg/ml) to the 1st well of 96 deep well plate
Add 200-300µL of the sample (lysate) from the 1.5mL centrifuged tube to the 1st well of 96 deep well plate
Add 400µL of isopropanol to the 2nd well of 96 deep well plate
Add 300µL of 80% enthanol to the 3rd well of 96 deep well plate
Add 300µL of 80% enthanol to the 4th well of 96 deep well plate
Add 300µL of DDW to the 5th well of 96 deep well plate
Add 100µL of DEPC-treated water to the 6th well of 96 deep well plate
Put the prepared 96 deep well plate in the automated DNA extraction machine and select the BOMB protocol
After the extraction is done, collect 50-100µL of the eluted sample as the DNA template for downstream experiments
