NucleoZOL is a highly efficient RNA isolation reagent for extracting high-quality miRNA and total RNA from a variety of starting materials. These include cells, tissues, and liquids of human or animal origin, as well as plants, yeast, bacteria, viruses, and other sources. Isolate RNA using a simple, one-phase extraction with NucleoZOL instead of an inconvenient two-phase chloroform extraction with a competitor “Zol”, and minimize the risk of sample loss and phenol carryover. The isolated RNA is ready for use in qRT-PCR, microarrays, RNase protection assays, poly A⁺ isolation, blotting, and other applications. NucleoZOL provides several advantages over a leading competitor “Zol” for rapid miRNA and total RNA isolation in terms of yield, quality, and purity.

The NucleoSpin RNA Set for NucleoZOL is a mini spin kit designed to combine NucleoZOL RNA extraction with NucleoSpin RNA purification. The kit contains silica membrane-based columns and the required buffers. After a sample is lysed in NucleoZOL (not provided in the mini spin kit), and contaminating DNA and proteins are pelleted, the supernatant is mixed with an optimized binding buffer for efficient binding of miRNA and large RNA to the silica membrane. A convenient bind-wash-elute procedure allows highly efficient isolation of pure RNA (miRNA and large RNA).

We compared NucleoZOL to the leading “Zol” two-phase extraction method. NucleoZOL resulted similar or greater yields with a far simpler protocol. RNA was extracted from a variety of starting materials and quantified using qRT-PCR.

Total RNA was isolated using NucleoZOL from 60 mg mouse heart tissue and analyzed on an Agilent Bioanalyzer. Total RNA was isolated with NucleoZOL in combination with the NucleoSpin RNA Set for NucleoZOL (from 5 x 10⁶ HeLa cells). RNA was analyzed on an Agilent Bioanalyzer; a RIN of 9 indicates high RNA quality.

Total RNA was isolated from 60 mg rat lung tissue using NucleoZOL and a competitor “Zol” product. DNA contamination was quantified by qPCR (competitor “Zol”=100%). Compared to a standard two-phase extraction, only a minimal amount of DNA is carried over during purification with NucleoZOL (Panel A). The A₂₆₀/A₂₃₀ absorption ratio of total RNA isolated using NucleoZOL shows excellent purity, while the lower A₂₆₀/A₂₃₀ ratio of total RNA isolated using the competitor “Zol” indicates phenol carryover (Panel B).

The NucleoZOL procedure is far easier than other “Zol” products because chloroform two-phase separation is not necessary. Cells and tissues are lysed and homogenized directly in NucleoZOL reagent, eliminating any chance of RNA degradation. Then contaminating molecules such as DNA, polysaccharides, and proteins are precipitated by the addition of water and removed by centrifugation. The NucleoZOL procedure allows the separate isolation of small and large RNA by adding ethanol and isopropanol, respectively. RNA can be reconstituted by RNase-free water.

NucleoZOL simplifies RNA isolation with a one-phase extraction

NucleoZol			Competitor Zol
	Sample homogenization			Sample homogenization
	Addition of water			Addition of chloroform
		Non-toxic			Toxic
	Centrifugation at RT			Centrifugation at 4°C
	Precipitation of contaminants			Liquid-liquid phase separation
	Aspiration of whole supernatant with RNA			Aspiration of aqueous phase with RNA
		Easy sampling of RNA			Inconvenient pipetting for phase separation
					Risk of carry-over of interphase/polar phase
					Refrigerated centrifuge neccessary
	Centrifugation at RT			Centrifugation at 4°C
	Precipitation of RNA			Precipitation of RNA
	Washing of RNA			Washing of RNA
	RNA in pellet			RNA in pellet
	Resuspension of RNA			Drying and resuspension of RNA
		No drying of RNA neccessary			Time-consuming drying of RNA
		Quick and easy

Advantages of NucleoZOL compared to a competitor “Zol”