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70
RNeasy Mini Handbook 10/2019
Troubleshooting Guide
This troubleshooting guide may be helpful in solving any problems that may arise. For more
information, see also the Frequently Asked Questions page at our Technical Support Center:
www.qiagen.com/FAQ/FAQList.aspx. The scientists in QIAGEN Technical Services are
always happy to answer any questions you may have about either the information and/or
protocols in this handbook or sample and assay technologies (for contact information, visit
www.qiagen.com).
Issue
Comments and suggestions
Clogged RNeasy spin column
a) Inefficient disruption
and/or homogenization
See “Disrupting and homogenizing starting material” (page 22) for details on
disruption and homogenization methods. Increase
g
-force and centrifugation time if
necessary.
In subsequent preparations, reduce the amount of starting material (see protocols)
and/or increase the volume of lysis buffer and the homogenization time.
If working with tissues rich in proteins, we recommend using the RNeasy Fibrous
Tissue Mini Kit (see page 91 for ordering information).
b) Too much starting
material
In subsequent preparations, reduce the amount of starting material. It is essential to
use the correct amount of starting material (see protocols).
c) Centrifugation before
adding ethanol not
performed (protocols for
tissues and mechanical
disruption of yeast)
Centrifuge the lysate before adding ethanol, and use only this supernatant in
subsequent steps (see protocols). Pellets contain cell debris that can clog the RNeasy
spin column.
d) Centrifugation
temperature too low
The centrifugation temperature should be 20–25°C. Some centrifuges may cool to
below 20°C even when set at 20°C. This can cause formation of precipitates that
can clog the RNeasy spin column. If this happens, set the centrifugation temperature
to 25°C. Warm the ethanol-containing lysate to 37°C before transferring it to the
RNeasy spin column.
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