Description
The EZBioscience® EZ-press Whole Blood RNA Purification Kit integrates phenol/guanidine-based sample lysis and silica-membrane purification of high-quality total RNA from fresh or frozen anti-coagulated blood. Whole blood samples are first lysed and homogenized in a strong denaturant and phenol containing buffer, which immediately inactivates RNases to ensure isolation of intact RNA. After addition of chloroform, the homogenate is separated into aqueous and organic phases by centrifugation. RNA partitions to the upper, aqueous phase while DNA partitions to the interphase and proteins to the lower, organic phase or the interphase. The upper, aqueous phase is collected and passed through the DNA Removing Spin Column which traps the DNA on the column, while RNA can pass through the column and retain in the flow-through. Ethanol is added to the flow-through, creating conditions that promote selective binding of RNA to the silica membrane. Then the sample is applied to a Spin Column for RNA, where the RNA binds to the membrane. Impurities are effectively removed by subsequent washing. Finally, high-quality RNA is eluted in Elution Buffer. The purified total RNA is suitable for use in a variety of downstream applications, including: RT-PCR, RT-qPCR, Northern blotting, nuclease protection assays, cDNA library preparation after poly (A)+ selection, and so on.
Components
Components | B0006 (100 Preps) |
RCL Buffer | 60 ml |
Lysis Buffer | 55 ml |
Wash Buffer* | 13 ml |
Elution Buffer | 10 ml |
DNA Removing Spin Columns (with Collection Tubes) | 100 preps |
Spin Columns for RNA (with Collection Tubes) | 100 preps |
*Before using for the first time, add 52 ml of 100% ethanol to the Wash Buffer.
Storage
Store the RCL Buffer and Lysis Buffer at 2 ~ 8°C upon reception, protect from light. Store other components at room temperature (Divide the Elution Buffer into 3 aliquots upon reception and store at room temperature). When using these buffers, be careful to avoid of contamination.
Experimental Procedure at a Glance
Features
1. Easy-to-follow protocol for lysis and homogenization.
2. High yields of RNA from fresh or frozen anti-coagulated blood.
3. High-quality RNA for all downstream applications: RT-PCR, RT-qPCR, Northern blotting, RNA-seq, Poly A+ RNA, and so on.
Representative Experimental Results
Figure 1. RNA isolated from 100 μl and 200 μl whole blood using EZBioscience® EZ-press Whole Blood RNA Purification Kit. RNA was eluted by 50 μl Elution Buffer, and 5 μl was loaded each lane. M: 250bp DNA Ladder; Lane 1: RNA isolated from 100 μl whole blood; Lane 2: RNA isolated from 200 μl whole blood.
For detailed information, please check the product manual.
