Sample Homogenization and Cell Lysis

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Complete the following steps to perform sample homogenization and cell lysis, the second step to extract DNA before library preparation.

Always cap and seal tubes before centrifuge and homogenize steps.

• Internal Control (T7 Phage Working stock at 1.09E+10 copies/ml)
• BashingBead Buffer
• ZR BashingBead Lysis Tube

Save the following program (total running time is ~20 minutes) on the bead beating system (homogenizer):

1. Homogenize at 6m/sec for 1 minute.
2. Rest for 5 minutes.
3. Homogenize at 6m/sec for 1 minute.
4. Rest for 5 minutes.
5. Homogenize at 6m/sec for 1 minute.
6. Rest for 5 minutes.
7. Homogenize at 6m/sec for 1 minute.
1. Combine the following volumes to prepare the Extraction Reagent Mix. Multiply each volume by the number of samples.

Reagent overage is not included. We recommend making one extra Extraction Reagent Mix to compensate for pipetting variation.

• Internal control T7 Phage (2 µl)
• BashingBead Buffer (400 µl)
2. For each sample (including external controls), add 402 µl Extraction Reagent Mix into a new BashingBead Lysis Tube.
3. Add approximately 400 µl bead suspension from step 6 of Urine Conditioning Buffer (UCB) Treatment to each BashingBead Lysis Tube.
4. Cap the BashingBead Lysis Tube.
5. Place BashingBead Lysis tubes on the homogenizer and run the saved program.
6. Centrifuge the BashingBead Lysis tubes at 10,000 × g for 2 minutes.