EXTRACTME PLASMID MIDI (EM16)

The plasmid DNA purification procedure utilizes pre-packed anion-exchange resin columns which efficiently and selectively bind nucleic acids. In the first isolation step, the pDNA is released from bacterial cells by alkaline lysis. Then the lysate is neutralized and all the cell residues along with the proteins and genomic DNA are separated in the centrifugation step. This alkaline lysis method and RNase treatment are used to obtain clear cell lysate with minimal genomic DNA/RNA contaminants. In the next step the lysate is applied to the purification column with the equilibrated resin and the DNA is bound. The washing stage effectively removes impurities and enzyme inhibitors. A suitable buffer with a high ionic strength allows the elution of the plasmid DNA, which is then concentrated and desalted by precipitation. The purified plasmid DNA may be used directly in all downstream applications such as PCR, qPCR, transfection, microinjection, Southern blotting, DNA sequencing, enzymatic restriction and so forth, or stored until ready to use.

PRODUCT SPECIFICATIONS

SAMPLE MATERIAL
Bacterial broth culture:
50-150 ml (high-copy number plasmids)
100-300 ml (medium- and low-copy number plasmids)

YIELD
200-600 μg of transfection-grade pDNA from 100 ml of cultured bacterial cells

TIME REQUIRED
70 min for EM16 and 100 min for EM17
additional ~60 min for DNA precipitation
DNA PURITY
A260/A280 ratio = 1.7 – 1.9

ENDOTOXIN REMOVAL (EM17)
<0.1 EU/μg verified by LAL

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