HiPure HY Plasmid EF Mega Kit

HiPure HY Plasmid EF Mega Kit is the core high-yield column product within the Magen endotoxin-free plasmid portfolio. Developed for high-copy plasmids, the kit processes up to 500 mL of bacterial culture and recovers up to 5 mg of plasmid DNA. Its cation-mediated workflow combines selective plasmid precipitation with filter-column purification. Because plasmid DNA is selectively concentrated before column capture, total recovery does not depend solely on the fixed adsorption capacity of a conventional silica membrane. The workflow supports transfection-grade plasmid DNA with endotoxin levels below 1 EU/µg and injection-grade DNA below 0.1 EU/µg.

Within the high-yield branch, P1116 serves as the core production-scale configuration. HiPure HY Plasmid EF 96 Kit (P1115) applies the high-yield principle to 96-well preparation, while P1117 extends processing to 1–1.5 L cultures with yields up to 10 mg and P1118 covers 10–15 L cultures with yields up to 100 mg. Laboratories processing medium- or low-copy plasmids, large constructs or a wider range of vector types should instead select HiPure Plasmid EF Maxi Kit (P1156).

Specifications

Engineering Characteristics

Ultra-high-capacity binding matrix

The modified binding matrix provides substantially greater plasmid-binding capacity than conventional silica-membrane columns, supporting high total recovery from high-copy plasmid cultures at mega scale.

Mega-scale plasmid preparation

Designed to produce up to 5 mg of low-endotoxin plasmid DNA per preparation, the workflow supports large-scale, transfection-oriented production requiring substantially higher output than routine maxi-prep formats.

Optimized for high-copy plasmids

The system is intended for high-copy cultures when total DNA output is the primary requirement. For medium- or low-copy plasmids, a salt-mediated EF workflow is generally more appropriate for controlling RNA background and maintaining reliable concentration measurements.

Technical Validation

The large-scale plasmid purification performance of the HiPure HY Plasmid EF Mega Kit (P1116) and HiPure HY Plasmid EF Giga Kit (P1117) was evaluated using plasmids ranging from approximately 3 kb to 12 kb and representing different culture productivity levels. P1116 was tested with 500 mL bacterial cultures, while P1117 was evaluated at the 1 L scale. Purified plasmid DNA was assessed by spectrophotometry, restriction digestion and Sanger sequencing.

Using P1116, total plasmid DNA yields ranged from 1.806 mg to 4.856 mg across four tested plasmids. DNA concentrations ranged from 0.722 to 1.942 µg/µL, with A260/A280 values of 1.91–1.94. The highest-yield culture produced approximately 4.9 mg of plasmid DNA, supporting the 5 mg preparation scale of the Mega workflow.

At the 1 L scale, P1117 produced 4.4–13.9 mg of plasmid DNA depending on the plasmid and culture productivity. DNA concentrations ranged from 0.489 to 1.539 µg/µL, with A260/A280 values of 1.91–1.92. The highest-yield plasmid culture exceeded the nominal 10 mg product range, while lower-producing plasmids generated 4.4–7.5 mg under the same culture-scale workflow.

When normalized to culture volume, both large-scale workflows generally retained at least 80% of the plasmid yield per millilitre obtained with the corresponding mini-prep reference. This indicates that increasing the culture volume from the mini-prep scale to 500 mL or 1 L did not result in a proportional loss of plasmid recovery.

Plasmid DNA purified with the large-scale workflow was successfully digested with EcoRI and HindIII across four plasmids of different sizes. The purified DNA was also evaluated by ABI 3730 Sanger sequencing and produced high-confidence sequence reads across all tested plasmids. These results support P1116 and P1117 as scalable high-yield purification routes for restriction digestion, sequencing and other molecular biology workflows requiring milligram quantities of plasmid DNA.

Kit Contents

Storage and Stability

The kit components can be stored dry at room temperature (15–25°C) and are stable for 24 months under these conditions. Buffer E2 and Buffer E4 may precipitate at low temperatures. Warm at 55°C until the precipitate is completely dissolved before use. After addition of RNase A, Buffer E1 is stable for 6 months when stored at 2–8°C.

For guidance on selecting between standard and low-endotoxin plasmid workflows based on copy number, culture input and processing format, please refer to the Plasmid DNA Kits Purchase Guide.