QIAGEN Plasmid Plus Midi Kit (25)

This QIAGEN kit is designed for fast and convenient purification of transfection-grade plasmid DNA with low endotoxin levels (<1 EU/µg), based on advanced silica technology (non-chaotrophic) and including a dedicated endotoxin reduction wash buffer. Endotoxin levels can vary depending on the host strain, culture media, and cell density, ranging from 1–3 EU/µg DNA in some cases. The midi format processes 25–50 ml culture volumes with a yield of up to 250 µg and a final DNA concentration of ≥1 µg/µl, handling 1–24 samples per run in approximately 20 minutes per run using parallel processing by centrifugation and vacuum. A vacuum manifold such as the QIAvac 24 Plus is recommended for use.

The bind-wash-elute procedure relies on unique binding chemistry in the QIAGEN Plasmid Plus columns. LyseBlue reagent is included for visual identification of optimum buffer mixing, and QIAfilter cartridges enable rapid lysate clearing. The purified plasmid DNA is obtained free of contaminants such as RNA, proteins and genomic DNA and is suitable for plasmids and cosmids used in transfection of sensitive cell lines, cloning, enzymatic restriction, in-vitro translation/transcription, and sequencing applications.

The product can be stored at room temperature for up to 24 months and requires the addition of RNase A and ethanol before use. The protocol uses Buffer P2 for lysis, followed by Buffer S3 and Buffer BB, a wash with 0.7 ml Buffer ETR, a further wash with 0.7 ml Buffer PE, centrifugation at 10,000 x g (9,700 rpm) for 1 min to remove residual wash buffer, and elution in 200 µl Buffer EB or water.