Introduction

Hipure Stool RNA Kit is specially designed for stool RNA extraction. This kit is suitable for extracting high-purity microbial or host cell RNA from ≤0.1g stool samples. The kit adopts silica gel column purification technology and original solution system, which can effectively remove humic acid and other inhibitory factors in stool samples. The purified RNA can be directly used in RT-PCR, Northern hybridization and other experiments.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA from 100-150mg stool sample
Applications	RT-PCR, Northern hybridization and other experiments
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Stool
Sample amount	100-150 mg
Elution volume	≥30μl
Time per run	≤50 minutes
Liquid carrying volume per column	100µg
Binding yield of column	800µl

Principle

The HiPure silica gel column uses a high binding ability glass fiber filter membrane as the substrate. Under the condition of high concentration of ionizing agent (such as Guanidinium chloride or guanidine isothiocyanate), the filter membrane can adsorb nucleic acid through hydrogen bonding and electrostatic and other physical factors, while protein or other impurities are not adsorbed and removed. The filter membrane that has adsorbed nucleic acids is washed to remove proteins and salts. Finally, low salt buffer solution (such as Buffer TE) or water can be used to wash out the nucleic acids adsorbed on the filter membrane. The obtained nucleic acid has high purity and can be directly used in various downstream experiments.

The stool samples are homogenized in the lysis solution, further lysed in a high-temperature water bath, and RNA is released into the lysis solution. Chloroform extraction removes genomic DNA and impurities, transfer the supernatant to an alcohol free binding solution, purify RNA through a column, and finally elute RNA with RNase Free Water. The purified RNA can be directly used for experiments such as PCR, Southern hybridization, and enzyme digestion.

Advantages

• High purity – unique adsorbent for more efficient removal of inhibitors
• High concentration – maximum extraction of total RNA from stool samples
• Sensitive – RNA can be purified at the level of PG

Kit Contents

Contents	R418502	R418503
Purification Times	50 Preps	250 Preps
HiPure RNA Mini Columns	50	250
2ml Collection Tubes	50	250
Glass Beads (0.1~0.6mm)	30 g	150 g
Buffer SPL	30 ml	140 ml
Buffer PHC	30 ml	140 ml
Buffer GRP	60 ml	250 ml
Buffer RW1	50 ml	250 ml
Buffer RW2 *	20 ml	2 x 50 ml
RNase Free Water	15 ml	30 ml

Storage and Stability

The kit components can be stored at room temperature (15–25°C) and are stable for 18 months under these conditions. At low temperatures, Buffer SPL may form precipitates, dissolve it by 55°C water bath. After receiving the product, Buffer PHC should be stored at 2-8°C.

Hipure Stool RNA Kit is specially designed for stool RNA extraction. This kit is suitable for extracting high-purity microbial or host cell RNA from ≤0.1g stool samples. The kit adopts silica gel column purification technology and original solution system, which can effectively remove humic acid and other inhibitory factors in stool samples. The purified RNA can be directly used in RT-PCR, Northern hybridization and other experiments.

Description

As this is a 2 gene kit, we recommend purchase of 2 of the accompanying RT-qPCR master mix reagent: oasig Lyophilised OneStep RT-qPCR Master Mix 150 reactions.

Norovirus is known to cause acute gastroenteritis. It is a small (27-38 nm), round, nonenveloped RNA virus belonging to the Caliciviridae family and is responsible for over 80% of non-bacterial outbreaks of gastroenteritis in the world. It affects individuals of all ages, with a distinct seasonal link to winter. It has a genome of 7.6 kb that is positive sense and has a single stranded linear confirmation. It encodes a major structural protein (VP1) of about 58 to 60 kDa and a minor capsid protein (VP2). Transmission occurs predominantly through ingestion of contaminated water, food and airborne transmission, as well as contact with contaminated surfaces. The ease with which norovirus is transmitted and the low infectious dose required to establish an infection results in extensive outbreaks in numerous environments, such as hospitals, hotels and schools. There is no antiviral drug available to treat this infection and little is known about its pathogenicity. However, it has been observed that the virus can be taken up by enterocytes where translation of viral nonstructural proteins can occur; it damages and alters intestinal microvilli, leaving them blunt and broadened, thus inhibiting absorption; it causes crypt cell hyperplasia and also leads to apoptosis of enterocyctes. An incubation period of 24-48 hours is usual. Infection is characterized by the acute onset of nausea, vomiting, abdominal cramps, aching limbs, raised temperature and diarrhoea that generally last for about 48 hours. However, more severe and prolonged infection may be observed in children and the elderly. There are five recognized norovirus genogroups, of which three (GI, GII, and GIV) are known to affect humans and, since 2002, variants of the GII.4 genotype have been the most common cause of norovirus outbreaks. There have been 31 different genotypes identified within the genogroups, with a wide degree of genetic variability present even within each genotype.

Exceptional value for money
Rapid detection of all clinically relevant subtypes
Positive copy number standard curve for quantification
Highly specific detection profile
High priming efficiency
Broad dynamic detection range (>6 logs)
Sensitive to < 100 copies of target Accurate controls to confirm findings

Amino-Tri-(Propargyl-PEG2-ethoxymethyl)-methane TFA Salt is a crosslinker consisting of an amino group with three propargyl groups. The amino group is reactive with carboxylic acids, activated NHS esters, carbonyls (ketone, aldehyde) etc. The propargyl groups can form triazole linkage with azide-bearing compounds or biomolecules via copper catalyzed Click Chemistry. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.

Amino-Tri-(Propargyl-PEG2-ethoxymethyl)-methane TFA Salt is a crosslinker consisting of an amino group with three propargyl groups. The amino group is reactive with carboxylic acids, activated NHS esters, carbonyls (ketone, aldehyde) etc. The propargyl groups can form triazole linkage with azide-bearing compounds or biomolecules via copper catalyzed Click Chemistry. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.