UM_TotalRNA_NucleoZOL

user's manuals about RNA or or

RNA isolationUser manualNucleoZOL

February 2016 / Rev. 02

user's manuals about RNA or or

Table of contents

1 Components

1.2 Reagents, consumables, and equipment to be supplied by the user 1.3 RNase-free work environment 1.4 About this user manual 2 Product description

2.1 The basic principle 1.1 Kit contents

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2.2 Product specifications 2.4 RNA reconstitution 4 Safety instructions 5 NucleoZOL protocols

2.3 Handling, preparation, and storage of starting materials 3 Storage conditions and preparation of working solutions

5.2 Isolation of total RNA 5.3

5.1 Isolation of small and large RNA in two separate fractions

NucleoZOL 18

2020

Isolation of total RNA in combination with NucleoSpin® RNA Set for

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6 Appendix

6.4 Product use restriction / warranty

6.3 Ordering information

6.2 Troubleshooting

6.1 Digestion of residual DNA solution

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user's manuals about RNA or or

1 Components

1.2 Reagents, consumables, and equipment to be supplied

by the user

Reagents

RNase-free water75 % ethanol70 % isopropanol100 % isopropanol

4-bromoanisole (optional, see p. 16)

1.5 mL, 2.0 mL or 15 mL centrifuge tubes (depending on the amount of sample to be processed per preparation) Sterile RNase-free tipsManual pipettorsVortex mixer

Centrifuge for microcentrifuge tubes

Equipment for sample disruption and homogenization Personal protection equipment (e.g., lab coat, gloves, goggles)Well ventilated working environmentRNase-free working environment

Consumables

Equipment

user's manuals about RNA or or

1.3 RNase-free work environment

The reagent has been tested for functionality. However, an RNase-free working environment is also a critical factor for performing successful RNA isolation and handling. Therefore, general recommendations to avoid RNase contamination should be followed:

Maintain a separate area, dedicated pipettors, and materials when working with RNA.

Wear gloves when handling RNA and reagents to avoid contact with skin, which is a source of RNases. Change gloves frequently.

Use sterile RNase-free plastic tubes. Tubes for lysate preparation and RNA precipitation have to be supplied by the user.

Keep all kit components sealed when not in use and all tubes tightly closed when possible.

1.4 About this user manual

Please read the detailed protocol if using NucleoZOL for the first time. Experienced users may refer to the short instruction manual.

All technical literature is available on the Internet at http://doc.guandang.net.

Please contact Technical Service regarding information about changes of to the current user manual compared to the previous revisions.

user's manuals about RNA or or

2 Product description

2.1 The basic principle

NucleoZOL is designed for the isolation of total RNA (small and large RNA) in a single fraction or in separate fractions from a variety of sample materials, such as cells, tissue, and liquids from human or animal origin, plants, yeast, bacteria, viral materials, and other sources.

One of the most important factors during the isolation of RNA is to prevent degradation. First, cells and tissues are lysed and homogenized in NucleoZOL reagent based on guanidinium thiocyanate and phenol. Contaminating molecules such as DNA, polysaccharides, and proteins are precipitated by the addition of water and removed by centrifugation. The NucleoZOL procedure allows the separate isolation of small and large RNA by adding ethanol and isopropanol, respectively. RNA can be reconstituted by RNase-free water. A chloroform-induced phase separation is not necessary for high-quality RNA isolation.

The RNA is ready for use in qRT-PCR, microarrays, RNase protection assays, poly A+ isolation, blotting, and other applications.

user's manuals about RNA or or

NucleoZOL

2.2 Product specificationsTable 1: Product specifications at a glance Technology Sample material (per mL NucleoZOL)7

One-phase extraction< 1 x 10 cultured cells, bacteria, and yeast,< 100 mg human/animal/plant tissue,< 0.4 mL (viral) fluids Small RNA (10–200 nt), large RNA (> 200 nt) Liver: 6–8 μg/mg tissue Kidney, spleen: 3–4 μg/mg tissue Muscle, brain, lung: 0.5–1.5 μg/mg Cultured cells: 4–10 μg/106 cells

Fragment size Typical yield (total RNA)

Typical yield (large RNA)

Liver: 5–7 μg/mg tissue Kidney, spleen: 3–4 μg/mg tissue Muscle, brain, lung: 0.5–1.5 μg/mg Cultured cells: 3–8 μg/106 cells

A260/280 (total RNA) Typical RIN (RNA integrity number) Elution volume

1.8–2.1> 9 flexible

MACHEREY-NAGEL– 02/2016, Rev. 02

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user's manuals about RNA or or

2.3 Handling, preparation, and storage of starting materials

Sample harvest and RNase inhibition

RNA is not protected against digestion until the sample material is flash frozen or disrupted in the presence of RNase inhibiting or denaturing agents. Sample harvest methods:

Use freshly harvested sample for immediate lysis and RNA purification.Samples can be stored in NucleoZOL after disruption at -20 °C to -70 °C for up to one year, at 4 °C for up to 24 hours, or up to several hours at room temperature. Frozen samples in NucleoZOL should be thawed slowly before starting with the isolation of RNA.

Flash freeze sample in liquid N2 immediately upon harvest and store at -70 °C. Frozen samples are stable up to 6 months. Mortar and pestle can be used to pulverize the sample in a frozen state. …… 此处隐藏：16806字，全部文档内容请下载后查看。喜欢就下载吧 ……