- DNA Purification Kit - For purification
of PCR products and DNA fragments from agarose gels.
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- Description
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- 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%.
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- Protocol
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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.
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- Storage
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- 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.
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- Applications
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- purification of DNA fragments, high-molecular-weight DNA and radiolabeled
probes.
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Quality control
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spectrophotometric control of purified DNA, cloning efficiency after
purification.
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References
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1. Boyle, J.S., Lew, A.M. (1995) TIG 11, 8.
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- Catalog #
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- K2
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Pack size
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100 preps
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