miRNeasy Kit for miRNA Purification

• Effective microRNA (miRNA) and total RNA purification, even from small samples
• Efficient enrichment of miRNA and RNAs <200 nucleotides
• Protocols for copurification or isolation of separate fractions
• High-purity RNA suitable for all downstream applications
• Automatable protocols and high-throughput processing in 96-well format

miRNeasy Kits enable purification of total RNA from all types of animal tissues and cells, including difficult-to-lyse tissues and small tissue and cell samples. Alternatively, an miRNA-enriched fraction and a total RNA (>200 nt) fraction can be purified separately (for separate purification, an RNeasy MinElute Cleanup Kit is required when using the miRNeasy Micro or Mini Kit and an additional RNeasy 96 plate when using the miRNeasy 96 Kit). Purification of 1–12 samples can be automated on the QIAcube Connect. High-throughput processing in convenient 96-well format can be performed using centrifugation or vacuum procedures.

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QIAzol based microRNA purification (miRNA purification) using miRNeasy Kits efficiently purifies RNA from small tissue (up to 5 mg tissue) and cell (up to 1 x 10⁶ cells) samples (see figure " High, reproducible recovery of miRNA from small cell numbers"). RNA can be purified from a variety of tissues and cells, including difficult-to-lyse tissues.

Purification of total RNA including miRNA allows direct comparison of miRNA expression levels with those of housekeeping reference genes or any other mRNA of interest. Alternatively, for some sensitive downstream applications, an miRNA-enriched fraction without larger RNA may be required. miRNeasy Kits enable efficient enrichment of RNA down to approximately 18 nt in size. The presence of very small RNAs is clearly visible after purification using an miRNeasy Kit, in contrast to samples prepared using an alternative kit (see figure " Effective enrichment of small RNA"). miRNeasy Kits efficiently purify RNA from both tissues and cells, even when low amounts of starting material are used (see figure " Effective purification from a range of starting amounts").

RNA can be purified with miRNeasy Kits from a variety of tissues and cells, including difficult-to-lyse tissues (see figure " Efficient copurification from a wide range of tissues"). miRNeasy procedures eliminate the possibility of contamination with salts or phenol which could interfere with later analyses (see figure " Highly pure RNA without phenol carryover"). In addition to higher purity, miRNeasy Kits offer superior yields to alternative methods of miRNA purification, such as using TRIzol Reagent (see figure " miRNeasy Mini Kit outperforms TRIzol").

RNA prepared using miRNeasy Kits is highly pure and ready for use in sensitive downstream applications.

miRNeasy Kits combine phenol/guanidine-based lysis of samples with silica membrane-based 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 tissues, to inhibit RNases, and also to remove most of the cellular DNA and proteins from the lysate by organic extraction.

miRNeasy Micro and Mini Kits are automatable on the QIAcube Connect. The miRNeasy 96 Kit enables vacuum and/or spin processing.

miRNeasy Kits enable purification of total RNA, which includes RNA from approximately 18 nucleotides (nt) upwards. Alternatively, an miRNA-enriched fraction and a total RNA (>200 nt) fraction can be purified separately (see flowchart " miRNeasy procedures"). Total RNA, including miRNA, is highly suitable for use in miRNA quantification by real-time RT-PCR. Enrichment of small RNAs in a separate fraction may be advantageous for certain applications where mRNA and rRNA could lead to increased background. This specialized protocol requires use of the RNeasy MinElute Cleanup Kit with the miRNeasy Micro or Mini Kit. If using the miRNeasy 96 Kit, 2 RNeasy 96 plates are required for every 96 samples.

Automated miRNA purification on the QIAcube Connect

Purification of total RNA, including miRNA, from the aqueous phase after phase separation can be fully automated on the QIAcube Connect using the miRNeasy Micro or Mini Kit (see figure " High, reproducible recovery of miRNA from small cell numbers"). The innovative QIAcube Connect uses advanced technology to process QIAGEN spin columns, enabling seamless integration of automated, low-throughput sample prep into your laboratory workflow. Sample preparation using the QIAcube Connect follows the same steps as the manual procedure (i.e., lyse, bind, wash, and elute), enabling you to continue using the miRNeasy Micro or Mini Kit for purification of high-quality RNA.

miRNeasy Kits allow purification of miRNA with total RNA for use in a variety of applications, including:

• RNA-seq
• Quantitative, real-time RT-PCR (e.g., using the miScript PCR System)
• Northern blot analysis
• Microarray analysis