Introduction

The Kit is designed for purification of total RNA, including miRNA and other small RNA molecules (18nt), from cultured cells and various animal and human tissues, including difficult-to-lyse tissues samples.

Details

Specifications

Features	Specifications
Main Functions	Isolation total RNA and miRNA from cell and tissue without MagZol reagent
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	Cell, Animal tissue, Plant tissue
Sample amount	Cells: ≤ 5 x 10^6, Animal tissue:<10mg
Elution volume	≥30μl
Time per run	≤40 minutes
Liquid carrying volume per column	800µl
Binding yield of column	100µg

Principle

Biological samples are first lysed and homogenized in a highly denaturing guanidine isothiocyanate-containing buffer, which immediately inactivates DNases and RNases to ensure isolation of intact DNA and RNA. The lysate is then passed through a Mini spin column. This column, in combination with the high-salt buffer, allows selective and efficient binding of genomic DNA. Flow-through from the column is digested by Proteinase K in the presence of ethanol. This optimized digestion, together with the subsequent addition of further ethanol, allows appropriate binding of total RNA, including miRNA, to the column. Contaminants are efficiently washed away and high-quality RNA is eluted.

Advantages

• High quality – one step RNA extraction reagent combined with silica gel column can obtain the highest concentration
• Fast – several samples can be extracted in 40 minutes
• High applicability – samples including animals, plants, bacteria, cells, etc.
• High concentration – efficiently remove macromolecular RNA, enrich small RNA and improve sensitivity
• Safe – no phenol chloroform extraction and no use of Trizol reagent

Kit Contents

Contents	R431102	R431103
Purification Times	50 Preps	250 Preps
HiPure RNA Mini Columns	100	2 x 250
2ml Collection Tubes	100	2 x 250
Proteinase K	48 mg	240 mg
Protease Dissolve Buffer	5 ml	15 ml
Buffer RLC	40 ml	200 ml
Buffer RWC	20 ml	80 ml
Buffer RW2*	20 ml	2 x 50 ml
RNase Free Water	10 ml	60 ml

Storage and Stability

Proteinase K should be stored at 2-8°C upon arrival. However, short-term storage (up to 12 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.

The Kit is designed for purification of total RNA, including miRNA and other small RNA molecules (18nt), from cultured cells and various animal and human tissues, including difficult-to-lyse tissues samples.

Overview

• Compatible with PAXgene Blood RNA Tubes
• Isolate true total RNA including siRNA and microRNA
• No phenol extractions
• Purify high-quality RNA in 30 minutes
• Purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen’s NGS services
• Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix

Norgen’s Preserved Blood RNA Purification Kit II provides a rapid method for the isolation and purification of total RNA from blood that has been preserved using PAXgene Blood RNA Tubes*. 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, without the use of phenol or chloroform.

Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. Norgen’s proprietary resin provides superior affinity to the full size range of RNA molecules, resulting in large and small RNA (miRNA) purification with better linearity and sensitivity. The purified RNA is of the highest integrity, and can be used in a number of downstream applications including real time RT-PCR, RT-PCR, Northern blotting, RNase protection and primer extension, expression profiling, miRNA cloning and amplification and Next Generation Sequencing.

* PAXgene Blood RNA Tubes are a product of PreAnalytiX GmbH.  PAXgene Blood RNA Tubes are not provided.

Details

Supporting Data

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Kit Specifications
Maximum Column Binding Capacity	50 μg
Maximum Column Loading Volume	650 μL
Size of RNA Purified	All sizes, including small RNA (< 200 nt)
Time to Complete 10 Purifications	30 minutes
Average Yield	5 – 20 μg per 2.5 mL preserved human blood

Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 1 year after the date of shipment. The DNase I should be stored at -20°C.

Component	Cat. 43500 (50 preps)
NPX1	2 x 110
NPX2	40 mL
NPX3	22 mL
NPX4	6 mL
NPX5	6 mL
DNase I	1 vial
Mini Spin Columns	50
Collection Tubes	50
Elution Tubes (1.7 mL)	50
Product Insert	1