N788
PHENOL-FREE TOTAL RNA PURIFICATION KIT

Grade:

CAS Number:
NA
Synonym(s):
RNA Purification Kit
Molecular Weight:
NA
Molecular Formula:
NA
Storage Condition:
RT
Risk Statements:
20/21/22, 36/37/38
Safety Statements:
36/37/39
Hazard Code:
None
UN Number:
NA
Code
Size
Price
Quantity
N788-KIT

KIT

$202.27

Overview

  • Purifies all sizes of RNA (from rRNA down to miRNA)
  • Purify RNA without phenol or chloroform
  • Purified RNA is suitable for all downstream applications

Detailed Description

Phenol-Free Total RNA Purification Kit allows purification of high quality RNA in just 20 minutes without the use of toxic chemicals. This system provides a rapid method for the isolation and purification of total RNA from cultured animal cells, tissue samples, blood, bacteria, yeast, fungi, plants and viruses. Lyse cellular or tissue sample, bind lysate to the column, wash column, and elute RNA. It's that simple. 

Click the Features tab to learn more about the Phenol-Free Total RNA Purification Kit.


Material Safety Data Sheet
Certificate of Analysis
Directions for Use

Phenol-Free Total RNA Purification Kit

  • Save time
    • Rapid and simple spin-column purification and concentration
    • Total RNA purification from multiple samples in 20 minutes
  • Reduce hazardous chemical use 
    • Eliminate phenol and chloroform
    •  Reduce interference by residual phenol in downstream applications including PCR
  •  Enhance recovery of total RNA including microRNA species
    • Efficient recovery from small sample sizes
    • Applicable to a wide range of sources

Purify all species and sizes of RNA without phenol/chloroform

AMRESCO's Phenol-Free Total RNA Purification Kit provides a rapid method for the isolation and purification of total RNA. In as little as 20 minutes, all sizes of RNA, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA), can be extracted without the use of phenol or chloroform.  The kit is an excellent tool for studies comparing expression of miRNA to other RNA species since both miRNA and mRNA can be analyzed from the same sample without further purification.  The kit can be used with samples from a wide range of organisms.

Enrichment of Small RNA Species

Figure 1. Size Range of RNA purified with Phenol-Free Total RNA Purification Kit versus a competitors kit. Total RNA was isolated from 1 x 109 E. coli cells with Phenol-Free Total RNA Purification Kit and a competitor’s kit. Five µL and 1 µL isolated RNA was analyzed on an agarose gel (Panel A) and the Agilent® 2100 BioAnalyzer RNA Nano 6000 chip (Panel B), respectively. Note the presence of small RNA species (red circle) in the samples isolated via Phenol-Free Total RNA Purification Kit and the absence of these RNA species in the competitor RNA preparation.

Amplification of miRNA and mRNA from the Same Extraction

Figure 2. Amplification of total RNA isolated from Phenol-Free Total RNA Purification Kit versus competitor kits. Blue: Amplification of RNA from Phenol-Free Total RNA Purification Kit; Red: Amplification of RNA from a competitor's phenol-based extraction reagent; Green: Amplification from competitor's spin-column technology.  RNA isolated from 0.75 million HeLa cells was polyadenylated according to Shi and Chiang (2005). Four microliters of the polyadenylated RNA were used in a 20 µL reverse transcription reaction with a poly T adaptor primer. One microliter of the reverse transcription was used in a 20 µL qPCR reaction with primers to the human microRNAs miR-19 and miR-21 and large mRNA S15). Amplification of RNA purified with Phenol-Free Total RNA Purification Kit matched amplification of both miRNAs and mRNA amplified with the phenol based extraction reagent. Amplification of microRNAs from the competitors spin column technology was not detectable.

Purification of Total RNA from Different Sources and Tissue Types

Figure 3. Total RNA was isolated from 5 x 108 E. coli cells, 1 x 106 HeLa cells, 100 µL rat blood and 10 mg hamster kidney. One microliter of the isolated RNA was analyzed on the Agilent® 2100 BioAnalyzer using an RNA Nano 6000 chip. RNA from a variety of sources extracted with Phenol-Free Total RNA Purification Kit consistently scored a RIN value between 8 and 10.

Sensitivity of Phenol-Free Total RNA Purification Kit

Figure 4. RNA Recovery by Phenol-Free Total RNA Purification Kit Approaches Single Cell Levels. Total RNA was extracted from serial dilutions of 293 HEK cells over a range of 1x106 cells to a single cell. The RNA was polyadenylated and reverse transcribed with an oligo dT primer. Three µL of the reverse transcription reaction was amplified by PCR  with primers to the human beta-actin gene. PCR products of beta-actin were detected from as little as a single cell. M is the marker lane.

Consistent Isolation of Large and Small RNA Species as a Function of Starting Amounts

Figure 5. Total RNA was isolated from 10 to 100,000 HeLa cells with Phenol-Free Total RNA Purification Kit (blue), a competitor’s spin-column kit (green) and a phenol-based RNA extraction method (red). Relative expression of miR-21 (Panel A), and S15 (Panel B) was determined by RT-qPCR of total RNA samples. One microliter of isolated RNA was  subjected to a 20 µL reverse transcription using miR-21 stem-loop reverse primer or oligo dT primer. Two microliters of the reverse transcription was added to a 20 µL real-time PCR reaction with primers to detect the human miR-21 (Panel A) and the S15 transcripts (Panel B). The resulting threshold cycle (Ct) values were plotted against input cell number. RNA isolated with Phenol-Free Total RNA Purification Kit had the best linearity (higher R2) and sensitivity (lower Ct) for both large RNA (S15) and small RNA (miR-21).

Consistent Detection of miRNA from Plasma

Figure 6. Isolation of Total RNA from Plasma by Phenol-Free Total RNA Purification Kit versus Competitors. Total RNA was isolated from 50, 100 or 200 µL of rat plasma in triplicates using Phenol-Free Total RNA Purification Kit (blue), a competitor spin column kit (green) and a phenol-based RNA extraction method (red). Two microliters of the  isolated RNA was subjected to a 20 µL reverse transcription with miR-21 stem-loop reverse primer or oligo dT primer. Three microliters of the reverse transcription reaction was transferred to a 20 µL real-time PCR reaction with primers to the human miR-21. Phenol-Free Total RNA Purification Kit is the only product that showed (1) consistent detection of miR-21 transcripts across all input volumes and (2) Ct values correlated to input volume (decrease Ct with increase input).

Consistent Performance of Diverse miRNA from Plasma in Downstream Applications

Figure 7. Total RNA including miRNA was isolated from 100 µL of mouse plasma in duplicate with Phenol-Free Total RNA Purification Kit, Competitor A spin-column Kit or a phenol-based RNA extraction method. One hundred nanograms of extracted RNA from each kit was applied to an Illumina microRNA expression profiling kit. Scatter plots display better consistency (better clustering) of replicate signals from AMRESCO 's samples. Genes with Pval < 0.01 for both replicates were in blue. The associated table suggested that AMRESCO's protocol recovered the same diversity of miRNA with better consistency (higher r2 value).

Enhanced Diversity of miRNA from Plasma

Figure 8: miRNA from plasma isolated with Phenol-Free Total RNA Purification Kit exhibits greater diversity than a leading competitor. Total RNA was isolated from 100 µL of plasma using AMRESCO's Phenol-Free Total RNA Purification Kit or 625 µL of plasma using Competitor A's miRNA Kit and was applied to an NCode expression profiling kit. Microarray images suggested that Phenol-Free Total RNA Purification Kit (left) achieves greater diversity of miRNA from smaller input amount of plasma than the competitor’s miRNA kit (right). Image courtesy of LC Sciences,Houston (www.lcsciences.com).

Kit Specifications


  Binding Capacity per Column   Up to 50 µg RNA
  Maximum Loading Volume Per Spin Column      650 µL
  Size of Purified RNA   All sizes, including < 200 nt     
  Time to Complete 10 Purifications             20 minutes
   RNA Yields  
  Tissue
 
Liver (10 mg)
 30 µg
Kidney (10 mg)
 10 µg
Brain(10 mg)
 12 µg
Blood (hamster, 100 µL )
 5 µg
Cells
Hela (1.0 x 106)
 15 µg
CHO (1.0 x 106)
 11 µg
Yeast (1.0 x 106)
 30 µg
E. Coli (1.0 x 106)
 43 µg

 

Related Products

Formaldehyde-Free RNA Gel Kit (N726)
  • Hood-free pouring and running - completely formaldehyde-free and glyoxal-free
  • Sample preparation - denature and stain samples in a single step
  • Instant band visualization - low background and brilliant green bands without the need for ethidium bromide
Agarose I™ (0710)
  • An all purpose agarose suitable for both analytical and preparative applications
RNA Markers