Cytoplasmic and nuclear RNA isolation kit

The Cytoplasmic and Nuclear RNA Purification Kit enables the convenient isolation of cytoplasmic and nuclear RNA from cultured cells and tissues.

Spin column

• 50 Preps
• 100 Preps

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Features and benefits

• Outstanding nuclear and cytoplasmic RNA separation and purification
• Easy and quick spin column format
• High RNA yield and quality
• Without phenol, isolate the entire range of RNA, including microRNA
• RT-PCR, qRT-PCR, RNA-Seq, arrays, and other applications can all use purified RNA
• Since cytoplasmic RNA is DNA-free, it can be used directly in RT-PCR and qRT-PCR
• Spin column chromatography, which makes use of Norgen's exclusive resin separation matrix, is the basis for purification

This kit offers a quick way to separate and purify nuclear and cytoplasmic RNA from small tissue samples and cultured animal cells.

Without the need for phenol, the kit can separate RNA of all sizes from the nuclear and cytoplasmic RNA fractions, including all small RNA species. The kit contains enough reagents to make 25 cytoplasmic and 25 nuclear RNA preparations or 50 cytoplasmic RNA preparations. It takes about 45 minutes to process 10 samples. There is also a 100-prep-size version of this kit available.

Background

In some situations, the ability to separate fractionated RNA rather than total RNA is preferable. For instance, in certain expression profiling investigations, isolating only mature cytoplasmic RNA might be better. As an alternative, nuclear RNA isolation might be preferable for research and analysis of pre-processed (non-spliced) RNA.

Since the cytoplasmic fraction has been demonstrated to be devoid of all traces of genomic DNA, this kit can isolate RNA for subsequent processes where DNA contamination must be avoided.

Details

Supporting data

Figure 1. Effective Separation of HeLa Cell Cytoplasmic & Nuclear RNA. Norgen's Cytoplasmic & Nuclear RNA Purification Kit was used to effectively separate cytoplasmic and nuclear RNA from 0.75 million HeLa cells in triplicate. Panel A: High quality cytoplasmic and nuclear RNA was purified using Norgen's kit. Note the integrity and quality of both cytoplasmic and nuclear RNA. Ten microliters of each 50 µL elution were run on a 1.5% formaldehyde-agarose gel. Lane M is Norgen's 1 kb RNA Ladder, lanes 1-3 contain nuclear RNA and lanes 4-6 contain cytoplasmic RNA. Panel B: Ten microliters of the above cytoplasmic and nuclear RNA isolated from HeLa cells using Norgen's kit was run on a 0.9% agarose DNA gel. Genomic DNA is clearly visible in the nuclear RNA fractions (lanes 1-3), however, no genomic DNA can be detected in the cytoplasmic RNA fractions (lanes 4-6). Note that an optional on-column DNase treatment protocol is provided to remove the genomic DNA in the nuclear fraction. Image Credit: Norgen Biotek Corporation

Figure 2. Superior Separation of HeLa Cell Cytoplasmic & Nuclear RNA. Norgen's Cytoplasmic & Nuclear RNA Purification Kit provides better separation of cytoplasmic and nuclear RNA from 0.8 million HeLa cells when compared to a leading competitor’s product. Panel A: Cytoplasmic and nuclear RNA purified from HeLa cells using Norgen's kit and a competitor's kit. Ten microliters of each 50 µL elution (Norgen or competitor's kit) of the cytoplasmic (C) or nuclear (N) RNA were run on a 1.5% formaldehyde-agarose gel. Higher yields of RNA with good integrity were isolated using Norgen's kit. Panel B: Ten microliters of the cytoplasmic and nuclear RNA isolated from HeLa cells using Norgen's kit and the competitor's kit was run on a 0.9% agarose gel. Genomic DNA only co-migrates with the nuclear fraction in RNA isolated using Norgen’s Cytoplasmic & Nuclear RNA Purification Kit, not the cytoplasmic fraction. Note that an optional on-column DNase treatment protocol is provided to remove the genomic DNA in the nuclear fraction. In contrast, significant genomic DNA contamination was observed in the cytoplasmic fraction of the RNA isolated using the competitor's kit. Image Credit: Norgen Biotek Corporation

Figure 3. Genomic DNA-free Cytoplasmic RNA. RT-PCR was performed using human beta actin primers on 2 µL of the 50 µL of cytoplasmic RNA isolated from 1 million HeLa cells using Norgen's Cytoplasmic & Nuclear RNA Purification Kit. Lane M is Norgen's PCR Sizer DNA ladder, Lanes 1 and 3 are the negative control (PCR only, without reverse transcript), and Lanes 2 and 4 are the actual RT-PCR that show the expected 166 bp RT-PCR product. The expected amplicon size from the gene copy is the same as that from the RNA transcript. The lack of product in lanes 1 and 3 indicates that no genomic DNA contamination is present in the isolated cytoplasmic RNA. All PCR products were resolved on a 1X TAE, 2% agarose gel. Image Credit: Norgen Biotek Corporation

Figure 4. Specific Separation and Subsequent Amplification of Cytoplasmic and Nuclear Transcripts. Norgen's Cytoplasmic & Nuclear RNA Purification Kit effectively separates cytoplasmic and nuclear RNA. RT-PCR was performed using human U2 snRNA (Panel A) or S14 (Panel B) primers on 0.5 µg of either cytoplasmic or nuclear RNA fractions isolated from 1 million HeLa cells using Norgen's Cytoplasmic & Nuclear RNA Purification Kit. In Panel A, an intense PCR product was observed only in the nuclear fraction when the nuclear-specific U2 snRNA primers were used. In Panel B, the majority of the PCR product for the housekeeping transcript of S14 was observed in the cytoplasmic fraction. The two results combined showed effective separation of the cytoplasmic and nuclear RNA using Norgen's Cytoplasmic & Nuclear RNA Purification Kit. All PCR products were resolved on a 1X TAE, 1.5% agarose DNA gel. Image Credit: Norgen Biotek Corporation

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Kit specifications

Source: Norgen Biotek Corporation

Storage conditions and product stability

Every solution needs to be stored at room temperature and kept tightly sealed. This kit remains stable for two years following the date of shipment.

Source: Norgen Biotek Corporation