Figure 1. Sequential Isolation of microRNA, Large RNA, Genomic DNA and Proteins from 1 x 10⁶ HeLa Cells.  Panels A and B show the separate isolation of large RNA and microRNA from 2 different samples of HeLa cells. Panel A is a 1X MOPS 1% agarose gel and Panel B is a 10% urea-PAGE gel. In both gels, Lane M is Norgen’s 1Kb RNA Ladder, Lanes 1 and 2 contain 3 µL out of the 50 µL elutions of the large RNA fraction, and Lanes 3 and 4 contain 3 µL out of the 50 µL elutions of the microRNA fractions. Panel C is a 1% agarose gel showing the gDNA isolated from the same 2 HeLa cell samples. Lane M is Norgen’s UltraRanger DNA Ladder and Lanes 1 and 2 contain 10 µL of each of the 100 µL elutions. Panel D is a 12% SDS-PAGE gel that contains the proteins that were isolated from the 2 HeLa cell samples. Lane M is a protein ladder and Lanes 1 and 2 contain 10 µL of the 100 µL elution of proteins that had been column purified. The large RNA, microRNA, gDNA and proteins are all intact and of the highest integrity and quality. Furthermore, the kit allows for the successful separate isolation of microRNA (<200 nt) and large RNA (>200nt).

Figure 2. Sequential Isolation of microRNA, Large RNA, Genomic DNA and Proteins from 5 x 10⁶ Yeast Cells.   Panels A shows the separate isolation of large RNA and microRNA from a plasmid-containing yeast culture. Panel A depicts the resolution of 1 µL of the 50 µL large RNA (Top) and small RNA (Bottom) fractions on an Agilent RNA Nano 6000 Chip. Panel B is a 1% agarose gel showing 10 µL of 100 µL of the gDNA and plasmid DNA isolated from the same yeast samples with Norgen’s UltraRanger DNA Ladder as molecular weight marker. Panel C is a 12% SDS-PAGE gel that contains 10 µL of 100 µL of the proteins that were isolated from the yeast samples. The large RNA, microRNA, gDNA and proteins are all intact and of the highest integrity and quality. Furthermore, the kit allows for the successful separate isolation of microRNA (<200 nt) and large RNA (>200nt) with the complete elimination of large rRNA in the small RNA fraction.

Figure 3. Isolate High Quality RNA.   RNA was isolated from 0.8 million HeLa cells using Norgen’s All-in-One Purification Kit and a leading market competitor for multiple analyte isolations. Half a microgram of the purified RNA (50 µL elution volume) was used as the template in a 20 µL reverse transcription reaction using oligo dT primers. One microliter of the RT reaction was used in a 20 µL qPCR reaction using the human GAPDH primers, and the results are shown in the PCR baseline graph. The blue lines correspond to the PCR results when RNA isolated using Norgen’s kit was used as the template, while the green lines correspond to the results when RNA isolated using the competitors kit was used as the template. From the graph it can be seen that Norgen’s kit isolated RNA with a greater sensitivity, and that more RNA was isolated from the same input as evidenced by the lower Ct values for the Norgen-isolated RNA.

Figure 4. Consistent Detection of Large mRNA, microRNA and Genomic DNA from the Same Sample.  Norgen’s All-in-One Purification Kit is the only kit available that concurrently isolates genomic DNA and all sizes of RNA including microRNAs without the use of phenol. RNA, DNA and proteins were isolated from 1 x 10⁶ of HeLa Cells using Norgen’s All-in-One Purification Kit in triplicate. One microgram of RNA was used in a 20 µL reverse transcription with oligo dT primer and miR-21 stem-loop primer. Three microliters of the RT reaction was used in 20 µL qPCR reactions for human S15 (for Large RNA) and miR-21 (for microRNA) genes, respectively. Also, 100 ng of gDNA was used in a 20 µL qPCR reaction for the 5S rRNA gene. All three genes studied showed efficient amplification within the expected range of Ct values. Moreover, as no phenol extraction and no splitting of samples was required, all replicates showed very similar Ct values suggesting that high consistency was maintained throughout the isolation of multiple biomolecules.