D. Biomek FX-Specific Pre-Run Recommendations

The Biomek FX automated platform allows users the flexibility to configure the robot’s deck configuration according to need. Because of this flexibility in deck configuration, it is likely that the deck used for writing a Biomek FX method will differ from an end-user’s deck. Therefore, it will be generally necessary to map an imported method onto an end-user’s deck configuration. To map an imported method onto your deck, please follow the instructions provided in the document Biomek FX
Deck Mapping (www.promega.com/automethods/beckman/biomek/default.asp).

E. Description of Automated MagneSil* ONE, Fixed Yield Blood Genomic System

This overview describes general liquid handling steps required for automated MagneSil* ONE, Fixed Yield Blood Genomic System and can be adapted to a variety of automated liquid handling robots. For additional information about adaptation to liquid handling robots other than those referenced above, please see Section VI. “General Guidelines for Adaptation to Alternative Robotic Platforms”.

1. Deck Preparation. Automated dispensing of all reagents into the appropriate plates. The use of plates instead of open reservoirs for reagent dispensing during the method decreases the probability of sample cross-contamination.Two hundred and ten microliters of Elution Buffer, Blood, 1,400µl of Lysis Buffer, Blood (with Anti-Foam added), 15µl of MagneSil® PMPs-Fixed Yield (after being resuspended on an orbital shaker for approximately 2 minutes), and 2,100µl of Alcohol Wash, Blood, are dispensed into seperate plates. Note: This deck preparation step can be separated from the rest of the purification method if desired. Plates can be predispensed, covered, and used at a later time.

2. Cell Lysis and DNA Binding. One hundred and ten microliters of Lysis Buffer, Blood, and 15µl of MagneSil* PMPs-Fixed Yield are premixed, then added to 60µl whole blood samples (stored in a 1.2ml deepwell round-bottom plate) and mixed well by pipetting. This step facilitates lysis of the blood cells and denatures protein and heme. An additional 430µl of Lysis Buffer, Blood, is added and mixed well by pipetting to promote efficient binding of the genomic DNA to the MagneSil* PMPs-Fixed Yield.
The sample plate is then placed onto the Deep Well MagnaBot* Device with the MagnaBot* 1/8 inch spacer. Samples are initially mixed on the Deep Well MagnaBot* Device to promote capture of any MagneSil* PMPs-Fixed Yield that have settled to the bottom of the well. After an additional 30 seconds of capture, the supernatant is discarded and the sample plate is moved off of the Deep Well
MagnaBot* Device.

3. Two Lysis Washes. Three hundred and sixty microliters of Lysis Buffer, Blood, is added to the samples and mixed well by pipetting. The sample plate is then placed onto the Deep Well MagnaBot* Device for 30 seconds to capture the MagneSil* PMPs-Fixed Yield.The supernatant is discarded and the sample plate is then moved off of the Deep Well MagnaBot* Device. This wash step is then repeated.

4. Three Alcohol Washes. Six hundred and eighty microliters of Alcohol Wash, Blood, is added to the samples and mixed well by pipetting. The sample plate is then placed onto the Deep Well MagnaBot* Device for 30 seconds to capture the MagneSil® PMPs-Fixed Yield.The supernatant is discarded and the sample plate is then moved off of the Deep Well MagnaBot* Device. This wash step is then repeated twice.

5. Drying/Removal of Residual Alcohol. The sample plate is then moved to the Heating/Cooling (H/C) ALP for 5 minutes. The Heating/Cooling ALP, with a recirculating water bath previously set at 80°C, is necessary for efficient sample drying (and elution).

6. Elution of Purified Genomic DNA. Two hundred and ten microliters of Elution Buffer, Blood, is added to the samples (still on the H/C ALP). The samples are incubated on the heating device with tip mixing intermittently for 8–10 minutes.The sample plate is then placed onto the Deep Well MagnaBot* Device for 1 minute to capture the MagneSil* PMPs–Fixed Yield.The supernatant is collected and saved in a clean 96-well round bottom plate.

7. Method Ends. Purified genomic DNA has been eluted into the 96-well round bottom plate.


F. General Guidelines for Adaptation to Alternative Robotic Platforms

We recommend the use of aerosol-resistant tips (barrier tips) for this method to decrease the chance of contaminating the instrument’s pipetting device. If your robotic platform uses fixed tips, be sure that the tips are washed thoroughly with bleach, hydrogen peroxide, or other appropriate wash solutions between pipetting steps to avoid sample cross-contamination. Also, if system liquid is used to perform pipetting steps, be sure to limit the exposure of samples to system liquid during all
pipetting steps by increasing the volume of leading air gaps that are used for pipetting. The MagneSil* PMPs–Fixed Yield used for this purification process settle quickily. We recommend thoroughly mixing the MagneSil* PMPs-Fixed Yield on the automated platform prior to dispensing to samples. Resuspension of the MagneSil* PMPs-Fixed Yield can be accomplished by thorough mixing or shaking. To efficiently elute the final purified samples from the MagneSil* PMPs-Fixed Yield,
it is important to include a heating step. This heating step ensures the samples are thoroughly dried following the last alcohol wash step and also helps to efficiently elute the final product from the MagneSil* PMPs-Fixed Yield.