Introduction
The EZBioscience® Tissue RNA Purification Kit PLUS integrates phenol/guanidine-based sample lysis and silica-membrane purification of high-quality total RNA from animal tissues and cells. Samples are first lysed and homogenized in Lysis 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 ethanol is added to provide appropriate binding conditions. The sample is then applied to the RNA Spin Column, where the total RNA binds to the membrane, and contaminants are effectively washed away. 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 | EZB-RN001-plus (100 Preps) |
Lysis Buffer | 55 ml |
Wash Buffer 1* | 13 ml |
Wash Buffer 2* | 13 ml |
Elution Buffer | 25 ml |
Spin Columns (with Collection Tubes) | 100 Preps |
*Before using for the first time, add 52 ml of 100% ethanol to the Wash Buffer 1 and Wash Buffer 2 separately.
Storage
Store the Lysis Buffer at 2 ~ 8°C, protect from light. Store other components at room temperature (When using these buffers, be careful to avoid of contamination). Dividing the Elution Buffer into small aliquots upon reception is suggested.
Experimental Procedure at a Glance
Features
1. Easy-to-follow protocol for lysis and homogenization.
2. High yields of RNA from animal tissues and cultured cells.
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 30 mg mouse muscle using EZBioscience® Tissue RNA Purification Kit Plus and TRIzol (eluted in 30 μl of Elution Buffer each, 5 μl RNA of each is loaded for electro- phoresis). M: 250bp DNA Ladder; Lanes 1 and 2: EZBioscience® Tissue RNA Purification Kit Plus; Lanes 3 and 4: TRIzol; These results showed that more RNA can be isolated from the same amount of tissue by EZBioscience® Tissue RNA Purification Kit Plus than TRIzol.
Figure 2. microRNA and lncRNA isolated from 293T cells are detected. These results showed that, this EZBioscience® Tissue RNA Purification Kit Plus can be used for microRNA and lncRNA purification and achieved good performance.
For detailed information, please check the product manual.
