Overview

• Purify all sizes of RNA (including microRNA) without the need for phenol
• Isolate and detect total RNA from 1 mL and up to 50 mL urine
• Provides high-quality RNA for sensitive applications – isolate RNA from as little as 100 cells
• Rapid processing time from sampling to downstream testing
• Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix

This kit provides a rapid spin column method for the isolation and purification of total RNA (including microRNA) from exfoliated cells that have been shed into the urine from the urinary tract. The kit purifies all sizes of RNA, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA). The RNA is preferentially purified from other cellular components such as proteins, as well as from the contaminating species found in urine such as glucose and salts, without the use of phenol or chloroform. The purified RNA is of the highest integrity and can be used in a number of downstream applications including real-time RT-PCR, qRT-PCR, Northern blotting, NGS, and expression array assays. Multiple samples can be processed in 20 minutes.

Background

RNA biomarkers from exfoliated cells can be used as non-invasive tools for a number of diagnostic and research applications including the diagnosis and monitoring of bladder, kidney, or other urinary-tract cancers. 

Details

Supporting Data

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Kit Specifications
Maximum Column Binding Capacity	50 μg
Volume of Urine Processed	1-50 mL
Maximum Input of Exfoliated Cells	1 x 106
Size of RNA Purified	All sizes, including small RNA (< 200 nt)
Time to Complete 10 Purifications	20 minutes (for Exfoliated Cells)30 minutes (for Bacteria)
Average Yield	~1μg RNA per 1 x 105 cells (For Exfoliated Cells – Varies due to cell density sample)~0.5μg RNA per 1 x 107 cells(For Bacteria – Varies due to cell density sample)

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.

Component	Cat. 22550 (25 preps)
Buffer SK	15 mL
Wash Solution A	18 mL
Elution Buffer E	6 mL
Micro Spin Columns	25
Collection Tubes	25
Elution Tubes (1.7 mL)	25
Product Insert	1

Introduction

The Kit provides fast purification of high-quality RNA from paxigene Blood RNA Tube using silica-membrane spin columns. RNA purified using the HiPure System is ready for applications such as RT-PCR, Northern blotting, poly A+ RNA (mRNA) purification, nuclease protection, and in vitro translation.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA from Paxgene RNA Tubes
Applications	RT-PCR, qPCR, Northern hybridization, NGS, nucleic acid protection, in vitro translation
Purification method	Mini spin column
Purification technology	Silica technology, DNA filtration technology
Process method	Manual (centrifugation or vacuum)
Sample type	Blood in the preservation tube
Sample amount	2.5ml
Elution volume	≥30μl
Liquid carrying volume per column	800µl
Binding yield of column	100μg

Principle

The Kit is for the purification of total RNA from 2.5 ml human whole blood collected in a PAXgene Blood RNA Tube. Purification begins with a centrifugation step to pellet nucleic acids in the PAXgene Blood RNA Tube. The pellet is washed and resuspended, and incubated in optimized buffers together with proteinase K. DNA wash The lysate is passed through a DNA Mini column. Ethanolis added to adjust binding conditions, and the lysate is applied to a column.RNA is selectively bound to the silica membrane as contaminants pass through. Remaining contaminants are removed in several efficient wash steps. Between the first and second wash steps, the membrane is treated with DNase I to remove trace amounts of bound DNA. After the wash steps, RNA is eluted in elution buffer and heat-denatured.

Advantages

• High quality – high purity total RNA can be directly used in various sensitive downstream applications
• Fast – isolation of several samples can be completed in 40 minutes by using column purification method
• Safety – no phenol chloroform extraction required
• Sensitive – direct lysis of blood, plasma and other samples without separation of leukocytes

Kit Contents

Contents	R416802	R416803
Purification Times	10 Preps	50 Preps
HiPure RNA Mini Columns I	10	50
gDNA Filter Mini Columns	10	50
2ml Collection Tubes	30	150
RNase Free Water	60 ml	250 ml
Buffer MBR1	10 ml	30 ml
Buffer MBR2	5 ml	15 ml
Buffer RW1	15 ml	60 ml
Buffer RW2*	6 ml	20 ml
Proteinase K	12 mg	50 mg
Protease Dissolve Buffer	1.8 ml	5 ml
DNase I	120 µl	600 µl
DNase Buffer	6 ml	30 ml

Storage and Stability

Proteinase K should be stored at 2–8°C upon arrival. DNase I should be stored at -20°C. However, short-term storage (DNase I up to 1 weeks, Proteinase K up to 8 weeks) at room temperature (15–25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under these conditions.

Experiment Data

The Kit provides fast purification of high-quality RNA from paxigene Blood RNA Tube using silica-membrane spin columns. RNA purified using the HiPure System is ready for applications such as RT-PCR, Northern blotting, poly A+ RNA (mRNA) purification, nuclease protection, and in vitro translation.

Propargyl-PEG8-bromide is a PEG reagent containing an alkyne group and a bromide group. The alkyne group can react with azide-bearing compounds or biomolecules in Click Chemistry reactions under the catalyzation of copper. The PEG spacer increases water-solubility of the molecule in aqueous media. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.

Propargyl-PEG8-bromide is a PEG reagent containing an alkyne group and a bromide group. The alkyne group can react with azide-bearing compounds or biomolecules in Click Chemistry reactions under the catalyzation of copper. The PEG spacer increases water-solubility of the molecule in aqueous media. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.