M.B. Enzymes GmbH

DNA Purification Kit - For purification of PCR products and DNA fragments from agarose gels.

Description

DNA purification kit consists of 3 components: melting solution, DNA binding suspension and wash concentrate. DNA molecules >20 bp and up to 100 kb length can be purified with a recovery of >80%.

Protocol

1. Add 23,75 ml deionized water and 25 ml ethanol to 1,25 ml wash concentrate to obtain 50 ml wash 1x solution.

2. Separate the DNA fragments by ectrophoresis in 0,5-1xTAE agarose gel. Do not use TBE because borate is not compatible with melting solution.

3. Excise the DNA fragment from agarose gel and add 300-400 µl of the melting solution. Apply 600 µl for 2% agarose. Incubate at 50-60°C with shaking for 5 min to melt agarose. Cool down. For purification of PCR fragments, mix 20-100 µl PCR reaction with 300-500 µl melting solition.

4. Add 5-15 µl binding suspension (will bind 1-4 µg DNA), mix and centrifuge 1 min at 6.000 rpm.

5. Remove supernatant. Add 500 µl of wash solution to the pellet and vortex. Repeat centrifugation. Wash twice.

6. Apply vacuum to dry the pellet.

7. Elute the DNA by resuspending the pellet in 50-100 µl deionized water. Incubate at 50-60°C for 5 min in shaker or vortex sometimes during incubation. Centrifuge 5 min at max. speed.

8. Transfer supernatant in a new tube. Repeat centrifugation step if necessary. Apply vacuum to concentrate the DNA.

Storage

Store melting solution in the dark at -20°C; DNA binding solution at +4°C and wash solution at room temperature. Vortex DNA binding solution vigorously every time before use.

Applications

purification of DNA fragments, high-molecular-weight DNA and radiolabeled probes.

Quality control

spectrophotometric control of purified DNA, cloning efficiency after purification.

References

1. Boyle, J.S., Lew, A.M. (1995) TIG 11, 8.

Catalog #

K2

Pack size

100 preps

DNA Purification Kit - For purification of PCR products and DNA fragments from agarose gels.
Description	DNA purification kit consists of 3 components: melting solution, DNA binding suspension and wash concentrate. DNA molecules >20 bp and up to 100 kb length can be purified with a recovery of >80%.
Protocol	1. Add 23,75 ml deionized water and 25 ml ethanol to 1,25 ml wash concentrate to obtain 50 ml wash 1x solution. 2. Separate the DNA fragments by ectrophoresis in 0,5-1xTAE agarose gel. Do not use TBE because borate is not compatible with melting solution. 3. Excise the DNA fragment from agarose gel and add 300-400 µl of the melting solution. Apply 600 µl for 2% agarose. Incubate at 50-60°C with shaking for 5 min to melt agarose. Cool down. For purification of PCR fragments, mix 20-100 µl PCR reaction with 300-500 µl melting solition. 4. Add 5-15 µl binding suspension (will bind 1-4 µg DNA), mix and centrifuge 1 min at 6.000 rpm. 5. Remove supernatant. Add 500 µl of wash solution to the pellet and vortex. Repeat centrifugation. Wash twice. 6. Apply vacuum to dry the pellet. 7. Elute the DNA by resuspending the pellet in 50-100 µl deionized water. Incubate at 50-60°C for 5 min in shaker or vortex sometimes during incubation. Centrifuge 5 min at max. speed. 8. Transfer supernatant in a new tube. Repeat centrifugation step if necessary. Apply vacuum to concentrate the DNA.
Storage	Store melting solution in the dark at -20°C; DNA binding solution at +4°C and wash solution at room temperature. Vortex DNA binding solution vigorously every time before use.
Applications	purification of DNA fragments, high-molecular-weight DNA and radiolabeled probes.
Quality control	spectrophotometric control of purified DNA, cloning efficiency after purification.
References	1. Boyle, J.S., Lew, A.M. (1995) TIG 11, 8.
Catalog #	K2
Pack size	100 preps