How can I improve recovery yields?

All of the protocols are designed to maximize yields for their respective kits.

The following tips are recommended:
A. Always use appropriate volume of buffer to sample. In general, using too little buffer can decrease yields.
B. Increase elution volumes. The minimum elution volumes for some of our kits (6ul for PCR Purification System, 35ul for Genomic DNA System) are designed to efficiently elute the purified product. Using higher volumes may increase total yields by up to 10%.
C. Pipette water or elution solution directly onto the column matrix. The BenevBio NucleoPur columns are designed to minimize any residual retention in the column, however pipetting directly onto the column matrix enables the water/elution solution to start releasing the purified product.

Can I increase the centrifugation speeds listed in the protocol?

Yes. Performing all centrifugation steps at top speed (~16,000 x g) will not adversely affect your yields in any way.

What is the minimum elution volume on the NucleoPur columns?

For the PCR Purification System, Gel DNA Extraction System, RNA Purification System, and DNA free RNA System the minimum elution volume is 6ul. However, one can always increase the elution volume to their desired preference (eg up 20ul).

For the Genomic DNA System, Total RNA Extraction System, and Viral RNA Extraction System the minimum elution volume is 35ul. However, one can increase the volume to their desired preference (eg. Up to 70ul).

What happens if I forget to add the Ethanol to the wash buffer?

First, add the appropriate volume of ethanol to the wash buffer concentrate.
Then simply re-purify the sample using a NucleoPur spin column and appropriate amount of buffer . For example if you are using the PCR Purification System and you mixed 100uls of PCR purification buffer to a 25ul PCR product and washed with 200ul wash buffer concentrate you would have a total volume of 325ul. Simply add the appropriate volume of PCR Purification Buffer (2 volumes in this case = 650ul) and spin through NucleoPur column and proceed with the wash step.

Why does my DNA or RNA sample float out of the well when loading it onto an agarose gel?

DNA samples purified using the PCR Purification System, Gel DNA Extraction System, etc. or RNA samples purified using the RNA Purification System, DNA free RNA System, etc. may float out of the wells due to carryover of residual ethanol from the final wash step.

Use either of the following steps to remove residual ethanol:
A. Re-purify the sample using a NucleoPur spin column.
B. Dry down the sample in a vacuum centrifuge, and resuspend the pellet in DNA/RNA free water.

How can I improve the ligation efficiency from DNA fragments extracted using the Gel DNA Extraction System?

Under certain conditions, enzymes from the ligation reaction may bind to NucleoPur particles affecting ligation efficiency. Centrifuge samples for 30 seconds and pipette the purified product into a new microcentrifuge tube.
Due to the chaotropic nature of the buffers used in the Gel DNA Extraction System some of the DNA may have denatured during the extraction process.

In order to promote renaturation of the single stranded DNA one can:
A. Elute the DNA from the NucleoPur column in 10mM Tris-EDTA buffer.
B. Set up the downstream reaction without the enzyme and incubate the mixture at 95C for 2 minutes to reanneal the single stranded DNA. Allow the mixture to cool to room temperature slowly before adding the enzyme and proceeding.

Can I use the PCR Purification System to purify enzymatic reactions such as digestions?

Yes. The PCR Purification System is designed to remove salts, enzymes, buffers, and nucleotides. Reactions that can be purified using the BenevBio PCR Purification System are restriction digests, ligase, priming, nick translation, and cDNA synthesis reactions.