SPINeasy® DNA/RNA/Protein All-In-One Kit

A: From the DNA extraction step, genomic DNA is extracted, which appears as high molecular weight band (>10,000bp) on the gel. Total RNA is extracted from the RNA extraction step, these typically appears as two ribosomal RNA bands and a low molecular weight bands corresponding to small RNAs.

A: Yes. Samples should ideally be fresh or freshly frozen and have not been frozen and thawed more than once.

A: SDS-PAGE (denaturing, rather than native PAGE) will be more appropriate as proteins would have been denatured in the extraction buffers and have lost their native conformations.

A: Yes. Theoretically when you are extracting genomic DNA (gDNA) using the kit, you get all type of DNA. However, test does not carry out to be certain. You can differentiate the gDNA from mtDNA by amplifying them using specific primer sequence.

A:

• As every protein has a different isoelectric point, there will be proteins that are not soluble at a certain pH. Adjust the pH of the resuspension buffer to suit the protein of interest.

• Increase the volume of resuspension buffer to allow more proteins to dissolve.

A: Yes, RIPA can be a buffer of choice to dissolve the protein pellet.

A: No, this kit is not suitable for ChIP applications as DNA-protein interactions are dissociated and extracted separately.

A: The kit has not been optimized for FFPE samples.

A: Total proteins are extracted from the protein extraction step, which should include membrane proteins. However, membrane proteins could be difficult to solubilize from the protein pellet. Thus, an appropriate resuspension buffer suitable for the particular protein may be required.

A: The kit has not been tested on brain samples.

A:

• Ensure the extraction was performed according to kit manual instructions.

• Sample with low nucleic acid content: (i) Increase amount of starting material; (ii) Elute in a smaller volume (50 µL).

• Insufficient lysis or over-lysis: Adjust FastPrep speed and/ or duration to achieve the optimal homogenization condition for the sample type. When using a vortex instead of a FastPrep for sample homogenization, lysis condition may be optimized by either reduced or extended vortexing duration.

• Poor sample quality: For best results, freshly prepared samples should be used.

• Poor elution: Ensure that Elution Buffer GD or Nuclease-free water is added to the center of the column membrane.

• RNase contamination: Work with nuclease-free tubes and pipette tips. Handle samples and perform all steps with clean gloves. Decontaminate work surfaces with RNase Erase® (Cat. No. 112440204).