Automation of the Macherey-Nagel NucleoSpin Robot-96 Plasmid Kit on the Zephyr Genomics Workstation IntroductionHere, we describe the use of the Zephyr Genomics Workstation for automated plasmid purification using The Macherey-Nagel NucleoSpin Robot-96 Plasmid Isolation Kit (CAT# 740 708.2). Using this system cells are lysed and purified plasmids are recovered in a two stage filtration, selective silica binding and elution under vacuum filtration. Special attention has been paid to enable a rapid parallel operation with strict timing control and perfectly uniform sample processing in order to get consistently high quality plasmid product. The results demonstrate expected yields of plasmid suitable for demanding downstream applications.

Uniform, High Quality Yields from a Parallel ProcessE.Coli DH5 Alpha harboring pGEM plasmids with a 359bp insert were grown in 1.3ml LB medium overnight. Cell pellets presented were introduced on the Zephyr Genomics Workstation for processing with the Macherey-Nagel NucleoSpin Robot-96 Plasmid Isolation Kit. All cultures were processed in parallel by arranging for stacked reagent containers and nested pipette tips.

Figure 1. Parallel Processing Enabled by Stacked Consumables.

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Figure 2. Consistent Yields of Purified DNA. Shown are mean recoveries and OD 260/280 ratios per column of a 96 well plate.

Figure 3. Plasmid DNA Integrity on Agarose Gel Electrophoresis. Visibly consistent plasmid is recovered from 96 wells with no evidence of low molecular weight species, indicating purity of plasmid.

Table 1. Time Savings vs. 8 Channel Protocols. In addition to processing uniformity time savings are achieved with respect to an alternate protocol.

ConclusionThe Zephyr Genomics Workstation can be used for automated plasmid purification using The Macherey-Nagel NucleoSpin Robot-96 Plasmid Isolation Kit (CAT# 740 708.2). Automation has been engineered to provide rapid parallel operation with strict timing control and perfectly uniform sample processing, achieving consistently high quality plasmid product. The results demonstrate expected yields of high quality plas-mid suitable for demanding downstream applications. 96 samples can be prepared in about one hour, and the Zephyr Genomics Workstation fully automates the process.

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Figure 4. Long Sequence Reads with Single Primer Extension Reaction. Shown is a typical electropherogram from a sequence reaction using purified plasmid. A quality score of 53 and a read length of 996 is obtained.

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Performance

Volume range: High Volume Head (HVH) 1 - 200 µL

Precision: High Volume Head (HVH) 1 - 5 µL: CV <5% 5 - 200 µL: CV <2% 2 - 25 µL or 50 µL: CV 5%

Deck Configuration

Caliper Life SciencesPart No. Description

125282 Zephyr Genomics Workstation with HV Head - 110V

125283 Zephyr Genomics Workstation with HV Head - 220V

119728 Optional Zephyr Environmental Enclosure

122000 LabChip GX Analyzer

760541 LabChip XT Instrument

Caliper’s Other Genomic Solutions

LabChip GX LabChip XT

Additional applications which can be automated on the Zephyr Genomics Workstation:• LabChip GX sample preparation for RNA quality assessment or DNA fragment analysis

• LabChip XT for fast, automated nucleic acid fractionation accurately and reproducibly

Zephyr’s small foot print fits on to a standard lab bench, requiring no special table, while offering flexibility for molecular biology applications: • DNA extraction • RNA extraction • Plasmid isolation • PCR purification • Sequencing reaction clean-up • PCR setup • Sequencing reaction setup • DNA and RNA normalization • Partial or whole plate

What’s included with your Zephyr Genomics Workstation: • PC controller and monitor • Vacuum filtration station and pump • Waste bottle • Plate/lid gripper • “PING!” Ultrasonic detector • Genomics Workstation GUI and Maestro software • Startup Kit