Description

MagExtractor -Plasmid- provides a simple and reliable method for the rapid purification of plasmid DNA from E. coli cells utilizing magnetic silica beads. This kit is based on binding properties of DNA to a silica surface in the presence of chaotropic agents (1). The purified plasmid can be used directly for automated fluorescent sequencing.

Features

  • Typical plasmid yield from an E. coli cell line carrying a high-copy plasmid is approx. 3-6 µg.
  • Suitable for high throughput extraction of plasmid from E. coli cells. The extraction time is 10-15 minutes.
  • Purified plasmid can be applied directly to sequencing, enzyme reaction, transformation, etc.
  • Does not contain hazardous substances, such as phenol or chloroform.

Applications

  • Plasmid purification

Storage condition

Store at 4ºC

Components

Binding Solution 130 ml x 2 (store at 4ºC or room temperature)
Magnetic Beads I   16 ml (store at 4ºC or room temperature)
Magnetic Beads II   16 ml (store at 4ºC or room temperature)
Suspension Solution   80 ml (store at 4ºC)
Lysis Solution I   80 ml (store at 4ºC or room temperature)
Lysis Solution II   20 ml (store at 4ºC)
Neutralization Solution   65 ml (store at 4ºC or room temperature)
Elution Solution   60 ml (store at 4ºC)
5x Loading Dye   11 ml (store at 4ºC)

Principle

The selectivity of extracted nucleic acids can be changed by optimization of the binding and washing solutions. MagExtractor -Genome- (Code No. NPK-101) extracts genomic DNA from various specimens (e.g. whole blood, cultured cells or animal tissues etc.). MagExtractor -RNA- (Code No. NPK-201) extracts total RNA from various specimens (e.g. cultured cells or animal tissues). MagExtractor -Plasmid- (Code No. NPK-301) extracts plasmids from E. coli cells, MagExtractor -Viral RNA- (Code No. NPK-401) is a kit for extracting viral RNA from serum or plasma specimens. MagExtractor -Plant Genome- (Code No. NPK-501) is a kit for extracting genomic DNA from various plant specimens (e.g., leaf, cultured cells, etc.). MagExtractor-PCR & Gel Clean up- (Code No. NPK-601) extracts DNA fragments from a PCR solution, enzyme solution, or agarose gel slices.

Application data

Example 1. Restriction enzyme digestion of the purified plasmid DNA

Plasmid DNA was purified from JM109 E. coli cells (from 1.2 ml cultured medium) carrying pUC18 by plasmid purification kits. Then, the purified plasmid DNA was digested by restriction enzymes. As shown in Fig.1, the purified plasmid DNA was digested completely.

Fig. 1. Electrophoretic analysis of the digested plasmid DNA

M: λ/HindIII Marker
U: Undigested
K: Kpn I digested
R: Rsa I digested

  Yield
(A260 nm)
Yield (A260 nm)
A260/
280 nm
A260/280 nm
MagExtractor 5.2 µg 1.85

Example 2. Sequencing analysis using the purified plasmid DNA

Sequencing analysis was performed using the purified pUC18 plasmid by MagExtractor -Plasmid-. As shown in Fig. 2, excellent quality data was obtained.

Fig. 1. Sequencing data (ABI PRISM 310 Genetic Analyzer)

References

  • B. Vogelstein and D. Gillespie, Proc. Natl. Acad. Sci. USA. 76: 615-619 (1979)