Description

MagExtractor -Genome- provides a simple and reliable method for the rapid purification of genomic DNA from various specimens (e.g. whole blood, cultured cells or animal tissues etc) using magnetic silica beads. This kit is based on binding properties of DNA to a silica surface in the presence of chaotropic agents (1)(2). The purified genomic DNA can be used directly for PCR experiments.

Features

  • Can be applied to extraction of DNA from whole blood specimens. A leukocyte or lymphocyte separation step is not necessary.
  • Does not contain hazardous substances, such as phenol or chloroform.
  • Purified DNA samples can be applied directly to other methods, such as PCR, etc.
  • Suitable for high-throughput extraction of genomic DNA from various specimens.

Applications

Sample Yield Purity(A260/280) Application
Blood 2 µg/100 µl whole blood 1.8 ± 0.1 PCR
Cultured cells 3 µg/5×10⁵ HeLa cells
Tissue* 5 µg/5 m;g tissue (swine testis)
Mouse tail* 3 µg/2 mm tail

*In the case of tissues or mouse tails, homogenization and/or lysis steps are necessary prior to extraction.

Storage condition

Store at 4ºC

Components

  (NPK-101) (NPK-102)
Lysis & Binding Solution 100 ml 500 ml
Washing Solution 200 ml 500 ml x 2
Magnetic Beads 5 ml 25 ml

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. Purification of genomic DNA from whole blood

Genomic DNA was purified from 100 ml whole blood specimens (human) using various purification kits. The purified genomic DNAs were analyzed by normal electrophoresis and pulse-field gel electrophoresis methods. As shown in Fig. 1, highly purified genomic DNA was successfully purified by MagExtractor -Genome- with high yield. Purified genomic DNAs from other company’s kits (company A and C) were estimated to contain proteins (assayed by absorbance at A260/280 nm (Table 1)). The average size of the purified genomic DNA by MagExtractor -Genome- was approximately 50 kb (Fig. 2).

Fig. 1. Electrophoretic analysis of purified genomic DNA from whole blood specimens

M: λ/Pst I Markers
1: MagExtractorTM
2: Company A
3: Company B
4: Company C


Fig. 2. Pulse-field gel electrophoresis analysis of the purified genomic DNA

Table 1. Comparison of yields and purity

  Yield
(A260 nm)
Yield (A260 nm)
A260/
280 nm
A260/280 nm
MagExtractor -Genome - 1.6 µg 1.80
Company A 3.5 µg 1.75
Company B 1.4 µg 1.81
Company C 2.9 µg 1.63

Example 2. Purification of genomic DNA from cultured cells

Genomic DNA was purified from HeLa cells (5×105 cells) using various purification kits. As shown in Fig. 1, highly pure genomic DNA was successfully purified by MagExtractor -Genome- with high yield. Purified genomic DNAs from other companies’ kits (company A, B and C) were estimated to contain RNA or proteins (Fig. 1, Table 1).

Fig. 1.Electrophoretic analysis of purified genomic DNA from cultured cells

M: λ/Pst I Markers
1: MagExtractorTM
2: Company A
3: Company B
4: Company C

Table 1. Comparison of yields and purity

  Yield
(A260 nm)
Yield (A260 nm)
A260/
280 nm
A260/280 nm
MagExtractor -Genome - 2.9 µg 1.84
Company A 8.1 µg 2.03
Company B 8.1 µg 1.82
Company C 9.7 µg 1.57

Example 3. PCR using purified genomic DNA samples

Genomic DNA was purified from 100 ml whole blood specimens from unrelated human donors using the MagExtractor kit. The yields and A260/280 nm (purity) were in the range of 1.4-2.2 µg and 1.83-1.84, respectively. PCR of the highly polytypic VNTR locus (MCT118) was performed using 1/20 volume of purified genomic DNA by Taq DNA polymerase. As shown in Fig. 2, distinct and highly polymorphic amplified bands were detected.

Fig. 1. Electrophoretic analysis of purified genomic DNA

M: λ/ Hind III Marker


Fig. 2. Analysis of the amplified products using purified genomic DNA

M: φx174 / Hinc II Marker
A-J: Samples
K: Negative control

Table 1. Comparison of yields and purity

  Yield
(A260 nm)
Yield (A260 nm)
A260/
280 nm
A260/280 nm
A 2.1 1.83
B 1.6 1.83
C 1.4 1.83
D 1.8 1.83
E 1.4 1.83
F 1.7 1.83
G 2.1 1.83
H 2.2 1.84
I 1.9 1.83
J 1.6 1.83

Example 4. PCR using purified genomic DNA samples

Genomic DNA was purified from 750 µl of supernatants of homogenized rat tissues in Lysis & binding solution. As shown in Fig.1, highly purified genomic DNA was successfully purified using MagExtractor -Genome-.

Fig. 1. Analysis of the purified genomic DNA from tissue samples

M: λ/ Hind I Markers
1: Rat Kidney 5mg
2: Rat heart 5mg

Table 1. Comparison of yields and purity

  Yield
(A260 nm)
Yield (A260 nm)
A260/
280 nm
A260/280 nm
Rat kidney 5mg 2.3 µg 1.86
Rat heart 5mg 1.6 µg 1.83

References

  • B. Vogelstein and D. Gillespie, Proc. Natl. Acad. Sci. USA. 76: 615-619 (1979)
  • R. Boom, C. J. A. Sol, M. M. M.Salimans, C. L. Wertheim-van Dillen, P. M. E. Dillen and J. van der Noordaa, J. Clin. Microbiol., 28: 495-503 (1990)