Product Manual
www.bioline.com/isolate
16
6. equipment anD reagents to Be supplieD By the user
•
Suitable container to hold sample
•
Mortar and pestle and liquid N
2
or rotor stator homogenizer
•
Microcentrifuge with rotor for 1.5ml and 2.0ml tubes
•
Shaking platform
•
70% and 96-100% ethanol
7. protocol
7.1 total rna isolation from plant tissue samples
Before you start:
•
Before using for the first time, add 96-100% ethanol to the Wash Buffers AR
and BR as indicated on the bottles and mix.
•
Avoid freezing and thawing of starting material.
•
If any of the buffers form precipitates upon storage, re-dissolve by gently
warming. Cool to room temperature before use.
•
For information on how to work with RNA, read Hints and Tips on page 19.
1. Homogenize and lyse up to 100mg of fresh or frozen plant tissue sample
using liquid nitrogen or a rotor-stator homogenizer.
Using liquid nitrogen
1.1. Grind the sample to a fine powder using a mortar and pestle in the presence of
liquid nitrogen. Take care that the sample does not thaw during or after grinding.
1.2. Transfer the sample to a 1.5ml microcentrifuge tube (not supplied).
1.3. Immediately add 450µl Lysis Buffer APR or BPR and homogenize the sample.
Proceed to next step. The sample can also be stored at this step at -20ºC.
Note: Most plant material can be lysed using Lysis Buffer APR and is the recommended
buffer for all applications. In case of low RNA yield, repeat the experiment with Lysis Buffer
BPR.
Using a rotor-stator homogenizer
1.1.
Transfer the sample to a suitable container.
1.2.
Add 450µl Lysis Buffer APR or BPR and homogenize the sample.
Note: Most plant material can be lysed using Lysis Buffer APR and is the recommended
buffer for all applications. In case of low RNA yield, repeat the experiment with Lysis Buffer
BPR.
1.3. Transfer the sample to a 1.5ml microcentrifuge tube (not supplied). Proceed to
next step. The sample can also be stored at this step at -20ºC.
