• iDi® 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 is the first choice for applications that rely on this property such as allele-specific PCR (asPCR) or allele-specific amplification (ASA).HiDi® Taq 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.Please note: This PCR mix is also available as a mix with a nuclease deficient variant, featuring higher robustness towards potential PCR inhibitors and compatibility with real-time dyes such as our GreenDye.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® Taq 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.HiDi® Taq DNA polymerase harbours a nuclease function and therefor is also suitable for use with hydrolysis probes (TaqMan® probes etc.). It has also been shown that HiDi® DNA polymerase family is highly suitable for quality control and mutation identification in CRISPR-Cas or TALEN-based applications.Several independently conducted studies show that HiDi® Taq 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 is the first choice for applications that rely on this property such as allele-specific PCR (asPCR) or allele-specific amplification (ASA).

Propargyl-PEG24-amine is a linker with 24 PEG units. The amine group is reactive with carboxylic acids, activated NHS esters, carbonyls (ketone, aldehyde) etc. The propargyl group is reactive with azide compounds under the catalyzation of copper. This linker has pretty good water-solubility due to its long PEG chain. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.

Propargyl-PEG24-amine is a linker with 24 PEG units. The amine group is reactive with carboxylic acids, activated NHS esters, carbonyls (ketone, aldehyde) etc. The propargyl group is reactive with azide compounds under the catalyzation of copper. This linker has pretty good water-solubility due to its long PEG chain. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.

Introduction

HiPure Bacterial RNA Kit uses silica gel column purification to simplify the extraction. The whole process does not require phenol chloroform extraction and time-consuming alcohol precipitation. The kit is suitable for efficiently extracting RNA from various bacterial samples. The purified RNA can be directly used for RT-PCR, Northern hybridization, etc. The kit has included lysozyme and glass beads, which can be used to treat gram-negative bacteria which is easy to be lysed, as well as gram-positive bacteria which is hard to be lysed, including enterococcus faecalis, staphylococcus, etc.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA from bacteria culture
Applications	RT-PCR, Northern blot, poly-A + purification, nucleic acid protection and in vitro translation
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Bacteria culture
Sample amount	Bacteria: <10^9
Elution volume	≥30μl
Time per run	≤40 minutes
Liquid carrying volume per column	800µl
Binding yield of column	100µg

Principle

Hipure silica gel column is based on glass fiber filter membrane with high binding force. Under the condition of high concentration of ionizing agent (such as guanidine hydrochloride or guanidine isothiocyanate), the filter membrane can adsorb nucleic acid through hydrogen bond and electrostatic, while protein and other impurities are not adsorbed and removed. The filter membrane adsorbed with nucleic acid is washed to remove the residual protein and salt. Finally, the nucleic acid adsorbed on the filter membrane can be washed out with low salt buffer (such as buffer TE) or water. The obtained nucleic acid has high purity and can be directly used in various downstream experiments.

Advantages

• Fast – several samples can be extracted in 30 minutes
• High purity – the purified RNA can be directly used in various downstream applications
• High recovery – RNA can be recovered at the level of PG
• Good repeatability – silica gel column purification technology can obtain ideal results every time
• Broad spectrum – it can deal with various bacteria, including Gram-positive bacteria that are difficult to be lysed
• Sufficient components – the kit contains carried lysozyme and glass beads

Kit Contents

Contents	R418101	R418102	R418103
Purification Times	10 Preps	50 Preps	250 Preps
gDNA Filter Mini Columns	10	50	250
HiPure RNA Mini Columns	10	50	250
2ml Collection Tubes	20	100	500
Glass Beads (0.1-0.6mm)	10 g	30 g	150 g
Plastic spoon	2	4	10
Lysozyme	20 mg	90 mg	400 mg
Protease Dissolve Buffer	1.8 ml	1.8 ml	10 ml
Buffer TE	1.8 ml	1.8 ml	5 ml
Buffer STL	5 ml	20 ml	90 ml
Buffer RLC	10 ml	30 ml	150 ml
Buffer RW1	10 ml	50 ml	250 ml
Buffer RW2*	5 ml	20 ml	2 x 50 ml
RNase Free Water	1.8 ml	10 ml	30 ml

Storage and Stability

The kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under these conditions. Due to the lack of antibacterial agents, RNase Free Water may be contaminated by bacterial or fungal when placed or operated at room temperature. It is recommended to pack and store at 2-8°C to reduce contamination.

HiPure Bacterial RNA Kit uses silica gel column purification to simplify the extraction. The whole process does not require phenol chloroform extraction and time-consuming alcohol precipitation. The kit is suitable for efficiently extracting RNA from various bacterial samples. The purified RNA can be directly used for RT-PCR, Northern hybridization, etc. The kit has included lysozyme and glass beads, which can be used to treat gram-negative bacteria which is easy to be lysed, as well as gram-positive bacteria which is hard to be lysed, including enterococcus faecalis, staphylococcus, etc.