FFPE RNA (including microRNA)/DNA Purification Kit

Supplier
Norgen Biotek
Cat. no.
1025000
Kit size
50
Links
Norgen Biotek Protocol Information sheet

Features

• High quality and integrity of the isolated nucleic acid - Both the purified total RNA and/or genomic DNA are of the highest quality and integrity, and can be used in any sensitive downstream application.
• Isolate a diversity of RNA species - All RNA species can be isolated, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA).
• High yields - Norgen’s FFPE RNA/DNA Purification Kit allows for the purification of high yields of total RNA or DNA.
• Rapid procedure - Isolate total RNA and/or genomic DNA from FFPE tissue sections using a rapid spin column format.
• Versatile procedure - The kit allows for the purification of either pure total RNA , pure genomic DNA or both RNA and genomic DNA through the use of tailored protease digestion times.

Norgen’s FFPE RNA/DNA Purification Kit provides a rapid method for the isolation and purification of total RNA (including microRNA) and genomic DNA from formalin-fixed paraffin-embedded (FFPE) tissue samples. Alternatively, the kit can be used to isolate total RNA alone or genomic DNA alone from FFPE tissue samples through varying the protease digestion time and performing optional RNase or DNAse digestions . Using formalin to fix tissues leads to crosslinking of the nucleic acids and proteins, and the process of embedding the tissue samples can also lead to fragmentation of the nucleic acids over time. Norgen’s FFPE RNA/DNA Purification Kit provides conditions that allow for the partial reversing of the formalin modifications, resulting in a high quality and yield of nucleic acids.

Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. The nucleic acids are preferentially purified from other cellular components without the use of phenol or chloroform. The process first involves deparaffinization of the FFPE samples through a series of xylene and ethanol washes. Next, the FFPE samples are digested with the provided Proteinase K and Digestion Buffer using an incubation time which is specific for the recovery of either RNA/DNA or DNA (please see the flow chart on page 4). Binding Solution and ethanol are then added to the lysate, and the solution is loaded onto a spin-column. Norgen’s resin binds nucleic acids in a manner that depends on ionic concentrations, thus only the RNA and/or DNA will bind to the column while the contaminants will be removed in the flowthrough or retained on the top of the resin. At this point, any traces of genomic DNA can be digested allowing for pure RNA samples to be isolated. Alternatively, traces of the RNA may be digested at this point if a pure sample of genomic DNA is required instead. The bound nucleic acid is then washed with the provided Wash Solution in order to remove any impurities, and the purified nucleic acid is eluted with the Elution Buffer.

Column Binding Capacity (RNA)	50 µg
Column Binding Capacity (DNA)	15 µg
Size of RNA purified	All sizes
Maximum Amount of Starting Material:	Paraffin Slices: 5 slices 20 µm thick
	Unsectioned Block: 25 mg

High Quality and Yield of Total RNA:

Total RNA was isolated from equal amounts of an FFPE kidney sample using Norgen’s FFPE RNA/DNA Purification Kit and a leading competitors kit. The purified RNA was then resolved on an Agilent BioAnalzyer. As it can be seen, Norgen not only isolated higher yields of total RNA, but the RNA was also of a higher quality as evidenced by the higher RIN values obtained with Norgen’s RNA.

High Quality Genomic DNA:

Genomic DNA was isolated from an FFPE kidney tissue sample using Nogen’s FFPE Nucleic Acid Purification Kit in duplicate and run on a 1.5% agarose gel. From observing the gel it can be seen that the kit isolates high yields of intact genomic DNA. Lane M is Norgen’s HighRanger 1kb DNA Ladder, and Lanes 1 and 2 contain the purified DNA samples.

• PCR & qPCR
• qRT-PCR
• primer extension
• expression array assays
• sequencing
• Southern blotting
• SNP analysis