For purification of total RNA from all types of tissue using gDNA Eliminator Solution
One-for-all tissue solution
Time-saving integration of RNeasy and QIAzol technologies
Fast non-enzymatic removal of genomic DNA
Pure RNA for use in downstream applications
RNeasy Plus Universal Kits integrate fast, convenient RNA purification with effective elimination of genomic DNA. Optimized protocols enable purification of high-quality RNA from any type of tissue, even difficult-to-lyse tissues. The kits include QIAzol Lysis Reagent for lysing fatty tissue and other types of tissue and RNeasy spin columns for purifying high-quality RNA. Kits are available in two formats: the RNeasy Plus Universal Mini Kit enables purification of RNA from up to 50 mg tissue and the RNeasy Plus Universal Midi Kit enables purification of RNA from up to 250 mg tissue. The RNeasy Plus Universal Mini Kit can be automated on the QIAcube.
For 10 RNA midipreps: RNeasy Midi Spin Columns, gDNA Eliminator Solution, Collection Tubes, RNase-Free Water and Buffers
RNeasy Plus Universal Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.
Effective removal of genomic DNA.
RNA was isolated from the indicated amounts of RNAlater stabilized rat kidney using the RNeasy Plus Universal Mini Kit, or was precipitated using a precipitation reagent from Supplier AII. RNA eluates were analyzed by real-time RT-PCR, with and without reverse transcription, using the QuantiFast Probe RT-PCR Kit and the Rotor-Gene Q.
High-quality RNA.
RNA was purified from rat tissue using the RNeasy Plus Universal Mini Kit, with or without gDNA Eliminator Solution. [A] Real-time RT-PCR analysis was carried out using RNA isolated from 25 mg frozen liver tissue. The CT value of all samples was comparable. DNA contamination was detected in the –RT control if gDNA Eliminator Solution was not used, whereas the samples isolated using gDNA Eliminator Solution showed minor or no DNA contamination. [B] RNA was purified from 25 mg stabilized heart tissues. Data obtained using the QIAxcel system demonstrates good RNA integrity regardless of whether RNA was isolated using gDNA Eliminator Solution.
RNeasy Plus Universal Mini and Midi procedures.
Performance
The RNeasy Plus Universal procedure allows high, reproducible RNA yields from any type of tissue, including fatty tissues and difficult-to-lyse tissue (see table). Yields can vary due to factors such as species and developmental stage, especially with adipose tissues.
Typical yields of total RNA with RNeasy Plus Universal Kits
Mouse/rat tissue (10 mg)
Yield of total RNA ((µg)
Adipose tissue
0.5–2.5
Brain
5–20
Heart
5–25
Intestine
10–60
Kidney
5–40
Liver
15–80
Lung
5–15
Muscle
5–35
Skin
2–5
Spleen
15–100
Analysis of purified RNA by real-time RT-PCR, and on the QIAxcel system, demonstrates high-quality RNA with good integrity (see figure "High-quality RNA"). gDNA Eliminator Solution and RNeasy technologies efficiently remove most of the genomic DNA contamination without DNase treatment (see figure "Effective removal of genomic DNA").
Principle
The RNeasy Plus Universal procedure enables purification of RNA from any type of tissue, including fatty or difficult-to-lyse tissues. RNeasy Plus Universal Kits integrate phenol/guanidine-based sample lysis and silica-membrane purification of total RNA. QIAzol Lysis Reagent, included in the kits, is a monophasic solution of phenol and guanidine thiocyanate, designed to facilitate lysis of all kinds of tissues and to inhibit RNases. The highly efficient lysis and the subsequent removal of contaminants by organic phase extraction enable the use of larger amounts of tissue with RNeasy spin columns. gDNA Eliminator solution effectively removes genomic DNA contamination and significantly improves separation of DNA into the interphase.
Procedure
The RNeasy Plus Universal procedure is easy to follow and, compared to phenol/chloroform treatment with a subsequent cleanup procedure, much faster and more efficient with better RNA purity and yield (see flowchart "RNeasy Plus Universal Mini and Midi procedures"). Total RNA is purified from up to 50 mg tissue using the RNeasy Plus Universal Mini Kit or up to 250 mg tissue using the RNeasy Plus Midi Kit.
Tissue samples are homogenized in QIAzol Lysis Reagent using the TissueRuptor, TissueLyser LT, or TissueLyser II. After addition of gDNA Eliminator Solution and chloroform, the homogenate is separated into aqueous and organic phases by centrifugation. The upper, aqueous phase with RNA is collected, mixed with ethanol, and RNA is purified using RNeasy spin columns. Total RNA binds to the spin column membrane, and phenol and other contaminants are efficiently washed away. High-quality RNA is eluted in RNase-free water.
The procedure provides an enrichment for mRNA since most RNAs <200 nucleotides (such as rRNAs and tRNAs) are excluded. The RNeasy Plus Universal procedure can be modified to allow the purification of total RNA containing small RNAs, such as miRNA, from cultured cells.Â
Applications
RNA purified using the RNeasy Plus Universal Kits is ideally suited for downstream applications that are sensitive to low amounts of DNA contamination, such as quantitative real-time RT-PCR. The purified RNA can also be used in other applications.
