Propargyl-PEG8-NHS ester

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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
• Detergent free
• Post-PCR Analysis – Enables post-PCR analysis

Applications

Ideal for contamination control in 

• PCR carry-over prevention
• RT-PCR
• RT-qPCR
• qPCR
• kPCR 
• RT-LAMP

Heat-labile Uracil-DNA Glycosylase

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.

Please refer to Protocol for Carry-over Contamination Control

Ordering Info

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.

• HiDi® stands for High Discrimination of mismatches at the 3’-terminus of primers in PCR. This myPOLS Biotec enzyme family is optimized for this feature and therefor, is the first choice for applications that rely on this property such as allele-specific PCR (asPCR) that is also termed allele-specific amplification (ASA).HiDi® 2x PCR Master Mix – ready to use mix simplifies your PCR setup. Only target-specific primers and template need to be added as the mix contains all components for a successful and reliable PCR. This ensures reproducible results, significantly reduces set-up times and the risk of pipetting errors.This PCR mix is also available with a full-length Taq DNA polymerase with a nuclease domain, featuring 100% compatibility with hydrolysis probes (TaqMan® probes etc.).Benchmarking with products of competitors conducted by us and others show that the HiDi® DNA polymerase family is the first choice for highly selective PCRs, such as genotyping by allele-specific PCR, HLA genotyping, analysis of single CpG methylation sites or the detection of mutations in a high background of wild-type sequences. By using HiDi® DNA polymerase, less than 10 copies of a mutation can be detected in a background of >10.000 wild-type copies straight away without any other tedious assay optimization.It has also been shown that HiDi® DNA polymerase family is highly suitable for quality control in CRISPR-Cas or TALEN-based applications.Several independently conducted studies show that HiDi® DNA polymerase is ideally suited for use in asPCR in numerous research areas ranging from mutation detection to genome editing. (read more)For research use and further manufacturing.In case you are aiming to use our RUO products as components or for your development of e.g. an IVD medical device, please contact us.

HiDi is available as:

HiDi® DNA Polymerase (>>)
HiDi® Taq DNA Polymerase (>>)
HiDi® 2x PCR Master Mix (>>)
HiDi® Taq 2x PCR Master Mix (>>)

Casestudies:
HiDi® DNA Polymerase: Applications from mutation detection to genome editing (read more)

Example Primer Design

Matching vs. mismatching nucleotide is placed at the 3′-end of the primer for best discrimination results.

Example Results – There´s no accounting for taste

Cilantro: some people love it in their food, some hate it. Here we are detecting a genomic SNP (rs72921001) in HeLa genomic DNA. This SNP is reported to be close to a number of genes coding for olfactory receptors. (Reference: Eriksson N. et al. (2012), “A genetic variant near olfactory receptor genes influences cilantro preference.”)

Considering, that only the C-allele specific primer is extended and yielding in a specific amplicon, we can conclude a genetic predisposition in disliking cilantro, as this SNP is significantly associated with detecting a soapy taste to cilantro.  

Allele-specific PCRs were performed from 1 ng/µl of HeLa gDNA in the presence of a realtime dye, indicating the amplification of the C-allele specific primer only. The A-allele specific primer is discriminated, thus not amplified up to 50 cycles.

PCR products were subsequently analysed on a 2.5% agarose gel. Specific product is visualized by ethidium bromide staining at the amplicon length of 109 bp.

HiDi® stands for High Discrimination of mismatches at the 3’-terminus of primers in PCR. This myPOLS Biotec enzyme family is optimized for this feature and therefor, is the first choice for applications that rely on this property such as allele-specific PCR (asPCR) that is also termed allele-specific amplification (ASA).

HiDi® 2x PCR Master Mix – ready to use mix simplifies your PCR setup. Only target-specific primers and template need to be added as the mix contains all components for a successful and reliable PCR. This ensures reproducible results, significantly reduces set-up times and the risk of pipetting errors.

Overview

DMSO resistant peelable heat sealing foil which is suitable for low temperature storage, high temperature uses and PCR testing.

• Heat sealing offers a 100% effective method of plate sealing, for complete seal integrity, as well as being quick and cost effective
• Our PeelASeal Foil Super is a laminate seal compatible with all plate types
• It can be removed by peeling, even with a plate which has been removed directly from -80°C storage
• PeelASeal Foil Super forms a complete seal to a plate enabling very low temperature uses, including very low temperature storage, and high temperature uses, such as PCR
• The seal demonstrates high solvent resistance (including DMSO) and can be utilized for short term compound storage at room temperature
• The seal is available as sheets, for use with manual and semi-automated sealers, such as our HeatASeal 500 Sealing Machine
• Also available in multiple roll formats compatible with specified automated heat sealers, such as our Wasp or Chameleon XT

IST-114 PeelASeal Foil SuperTM Heat Sealing Film