Science Catalog 2012 Life Cell Signaling PureYield™ Plasmid Midiprep System Product PureYield™ Plasmid Midiprep System Available Separately Cell Resuspension Solution (CRA) Cell Lysis Solution (CLA) Neutralization Solution (NSB) Eluator™ Vacuum Elution Device 315 ml A7115 117.00 315 ml A7125 117.00 500 ml A1485 80.00 4 each A1071 161.00 A1071 For Laboratory Use. A2492, A7115, A2495, A7125, A1485, A2496, A6740, A6742, A6744 For Research Use Only. Not for Use in Diagnostic Procedures. Description: The PureYield™ Plasmid Midiprep System is designed to isolate transfection-quality plasmid DNA. The system provides a rapid method for purification of 100–200μg of plasmid DNA from 50ml bacteria culture. Plasmid DNA can be purified in as little as 30 minutes with the vacuum protocol, greatly reducing the time spent on purification compared to silica resin or other membrane-column methods. An alternative protocol allows purification of over 400μg of high-copy-number plasmid from 250ml of E. coli culture. The PureYield™ Plasmid Midiprep System incorporates a unique Endotoxin Removal Wash designed to remove substantial amounts of protein, RNA and endotoxin contaminants from purified plasmid DNA. Removal of contaminants improves the robustness of sensitive applications such as eukaryotic transfection, in vitro transcription and coupled in vitro transcription/translation (e.g., TNT® Quick Coupled Transcription/Translation System). Purification is achieved without isopropanol precipitation of purified plasmid DNA or extensive centrifugation, providing rapid purification from a single method. The system has been designed for use with centrifugation or vacuum (e.g., the Vac-Man® Laboratory Vacuum Manifold). The Eluator™ Vacuum Elution Device is used to elute nucleic acids from PureYield™ Midiprep or Maxiprep columns. It consists of two pieces, a blue base and a clear column assembly. The base interfaces with a Vacuum Manifold that contains Luer-Lok® fittings, such as the Vac-Man® Laboratory Vacuum Manifold (Cat.# A7231), and holds a 1.5ml tube to capture the eluted nucleic acids. The column assembly accepts PureYield™ Midiprep or Maxiprep columns. The Eluator™ Device eliminates the requirement for a centrifuge with a swinging bucket rotor for nucleic acid purification, simplifying and speeding purification protocols. Features: • Improved Productivity: Vacuum protocol allows plasmid DNA purification in as little as 30 minutes. • Confidence in Results: High purity and concentration of plasmid DNA gives proven performance in transfection, in vitro expression and other molecular biology applications. • Ease of Use: Simple protocol eliminates tedious high-speed centrifugation, gravity-drip columns, and post-elution alcohol precipitation. • Flexibility: PureYield™ membrane column allows purification of large amounts of plasmid DNA, exceeding the capabilities of other midiprep systems. Storage Conditions: Store all system components at 22–25°C. Protocol PureYield™ Plasmid Midiprep System Technical Manual Part# TM253 PureYield™ Plasmid Midiprep System. Proper assembly of Lysate Clearing Column (blue) and DNA Binding Column (white) for use with the DNA Purification by Vacuum protocol. 1,000 2,000 3,000 4,000 5,000 6,000 7,000 8,000 0 Qiagen Eppendorf PureYield™ System DNA Preparation Methods Comparison of transfection of plasmid DNA purified with the PureYield™ System and other midiprep systems. psiCHECK™-2 Vector (Cat.# C8021), which carries a firefly luciferase gene, was isolated from E. coli using the PureYield™ System, the Qiagen HiSpeed® Plasmid Midi Kit or the Eppendorf Perfectprep® Plasmid Midi Kit. HeLa cells were transfected using 0.07μg of DNA in a total of 25μl. The firefly luciferase signal was monitored with the Dual-Glo™ Luciferase Assay System (Cat.# E2920). Size Cat.# Price (Fr) 25 preps A2492 253.00 100 preps A2495 886.00 300 preps A2496 2393.00 Midiprep PureYield™ Done in 30 Minutes Wizard® 99 Minutes Qiagen 200 Minutes Comparison of time required per midiprep for different systems. Each system protocol was performed according to the manufacturer’s instructions using 50ml of an overnight culture of JM109 bacteria transformed with highcopy-number plasmid [pGEM®-3Zf(+) Vector, Cat.# P2271]. Total time to perform midiprep is noted. 118 For complete and up-to-date product information visit: www.promega.com/catalog Luminescence (RLU) 4701MA 4594TA 4587MB Pagina 121
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