Cod Uracil-DNA Glycosylase (Cod UNG) from Atlantic Cod is the only commercially available UNG enzyme that is completely and irreversibly inactivated by moderate heat treatment. The enzyme is produced in a recombinant E. coli (ung-) strain that contains a modified Cod UNG gene.
Key Features:
Heat-labile – Completely and irreversibly inactivated at 55°C
Contamination control – ideal in applications below
Use of Cod UNG makes contamination control possible in RT-PCR
Does not degrade PCR product post-PCR. This makes downstream use of the PCR product possible
High purity enzyme, tested free of contaminating nucleases
There are several commercially available Uracil-DNA glycosylases on the market today. Most of them are of bacterial origin and work well if you have no intention to further analyze the PCR products post-PCR. However, if you want to store your PCR products for downstream analysis such as cloning and sequencing, the reactivation of UNG and subsequent degradation of your PCR products are a problem with most of the commercially available UNGs. Cod UNG from ArcticZymes is completely and irreversibly inactivated by heat thus ensuring that sample integrity is maintained long-term regardless of storage conditions.
This is illustrated in figure 1, below
Figures
Properties
Recommended Protocols
1. Contamination control in PCR, qPCR and one-step RT-qPCR
Cod UNG works in all commercially available master mixes.
Be sure that you have used dUTP containing dNTP mixes in your previous PCR experiments.
Add 0.2 U Cod UNG directly to your 20 µl PCR reaction.
pre-incubate for 5 min at room temperature.
For RT-qPCR, reverse transcribe your RNA at 50-55°C.
Run your PCR.
Store your PCR product at -20°C or 4°C degrees.
2. Contamination control in RT-LAMP
Cod UNG is ideal for contamination control in RT-LAMP. One unit of Cod UNG per 30 μl reaction is sufficient for removing even high concentrations of carry-over contamination.
Ensure that you use dNTP mixes containing dUTP in your experiments.
Check that the RT-LAMP reaction is compatible with dUTP by running side-by-side reactions containing different ratios of dUTP to dUTP (100% dUTP, 90% dUTP, 80% dUTP and 0% dUTP).
Add 1 U Cod UNG directly to your 30 µl RT-LAMP reaction.
Prepare the reaction mix on ice.
Analyze your RNA at 65°C, no preincubation is necessary.
Cod Uracil-DNA Glycosylase (Cod UNG) from Atlantic Cod is the only commercially available UNG enzyme that is completely and irreversibly inactivated by moderate heat treatment. The enzyme is produced in a recombinant E. coli (ung-) strain that contains a modified Cod UNG gene.
Isolate all sizes of circulating and exosomal RNA, including microRNA
Versatile plasma/serum input ranges
No phenol extractions
No carrier RNA
Bind and elute all RNA irrespective of size or GC content, without bias
Concentrate circulating RNA and exosomal RNA into a flexible elution volume
High quality, purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen’s NGS services
Compatible with Streck Cell-Free RNA BCT® Tubes
Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix
These kits provide a fast, reliable and convenient method to purify and concentrate high quality, high purity and inhibitor-free cell-free circulating and exosomal RNA using a convenient spin column method. These kits can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate. Plasma samples prepared from blood collected on heparin should not be used, as heparin can significantly interfere with many downstream applications such as RT-PCR. The purified plasma/serum RNA is fully compatible with all downstream applications including PCR, qPCR, methylation-sensitive reverse transcription qPCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, expression array assays, and NGS.
Background
Plasma/Serum cell-free circulating RNA or exosomal RNA has the potential to provide biomarkers for certain cancers and disease states. Exosomes are 40 – 150 nm membrane vesicles, which are secreted by most cell types. Exosomes can be found in saliva, blood, urine, amniotic fluid and malignant ascitic fluids, among other biological fluids. Evidence has been accumulating recently that these vesicles act as cellular messengers, conveying information to distant cells and tissues within the body. These exosomes may play a functional role in mediating adaptive immune responses to infectious agents and tumours, tissue repair, neural communication and transfer of pathogenic proteins. For this reason exosomal RNAs may serve as biomarkers for various diseases including cancer. As the RNA molecules encapsulated within exosomes are protected from degradation by RNAses they can be efficiently recovered from biological fluids, such as plasma or serum.
Plasma/Serum RNA Purification Mini Kit
This kit can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 50 µL to 200 µL. The purified plasma/serum RNA is eluted in a flexible final volume of 10 µL to 25 µL.
Plasma/Serum RNA Purification Midi Kit
This utilizes a two column method, and can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 250 µL to 1.5 mL. The first column will handle the large volume input of bodily fluids that is followed by a concentration on a mini column for a final elution of 50 µL to 100 µL.
Plasma/Serum RNA Purification Maxi Kit
This kit can purify RNA from fresh or frozen serum or plasma samples prepared from blood collected on either EDTA or Citrate, from volumes ranging from 2 mL to 5 mL. The first column will handle the large volume input of bodily fluids that is followed by a concentration on a mini column for a final elution of 50 µL to 100 µL.
All sizes, including miRNA and small RNA (<200 nt)
Elution Volume
50-100 μL
Time to Complete 10 Purifications
35-40 minutes
Average Yields*
Variable depending on specimen
*Please check page 7 of the Product Insert for Average Plasma/Serum Yields and Common RNA Quantification Methods.
This kit is suitable for the isolation of RNA from fresh or frozen serum or plasma prepared from blood collected on either EDTA or Citrate. Plasma samples prepared from blood collected on heparin should not be used as heparin can significantly interfere with many downstream applications such as RT-PCR.
Storage Conditions and Product Stability All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment. It is recommended to warm Lysis Buffer A for 20 minutes at 60°C if any salt precipitation is observed.
