Available Options:
PRINCIPLE
The EXTRACTME VIRAL RNA KIT utilizes spin minicolumns with membranes which efficiently and selectively bind nucleic acids at high concentration of chaotropic salts. During the first isolation step, the material is lysed under highly denaturing conditions to inactivate nucleases and to ensure isolation of intact viral RNA. RNA bounds to a Purification Column membrane by addition of alcohol. A three-step washing stage effectively removes impurities and enzyme inhibitors. Purified RNA is eluted with the use of low ionic strength buffer and may be used directly in all downstream applications, such as RT-PCR, RT-qPCR, cDNA synthesis.
PRODUCT SPECIFICATIONS
SAMPLE MATERIAL
swab in viral transport medium (e.g., nasopharyngeal swabs, mouth and throat swabs)
EFFICIENCY
dependent on the sample material amount and type
BINDING CAPACITY
~ 120 μg RNA
TIME REQUIRED
10–12 minutes
RNA PURITY
A260/A280 ratio = 1.9 – 2.1
EXTRACTME VIRAL RNA KIT – TROUBLESHOOTING
| Problem | Possible cause | Solution |
|---|---|---|
| RNA Purification Column becomes clogged during purification. | The purification column is overloaded. | Decrease the amount of a sample material. |
| Low RNA yield. | Sample material was incorrectly stored or preserved: RNA degradation. | Ensure that the swab was taken correctly and that the transport conditions were adequate. |
| Too little sample material was used. | Take more sample material. A proper amount of a sample is dependent on the kind of a material examined and needs to be optimized individually. | |
| The purification column has become clogged. | See “RNA Purification Column becomes clogged during purification”. | |
| RNases are present. | See “RNase elimination” in Manual section VIII. Recommendations and Important Notes. | |
| RNA is still bound to the column membrane. | Repeat the RNA elution. | |
| Low purified RNA concentration. | Too much of elution buffer was used. | Decrease the vREB volume up to 30 μl. |
| Too low A260/A280 ratio of purified RNA. | Remainings of buffers present in the eluate. | Ensure that the purification column had been properly dried before elution and no droplets remained on the ring. If necessary, increase centrifugation speed at step 7 of Isolation Protocol (section X) to 18 000 x g. Carefully remove the column from a collection tube. |
| Incomplete sample loading. | Make sure that the lysate has passed completely through the RNA Purification Column before proceeding through washing steps. If necessary, increase centrifugation speed at step 4 of Isolation Protocol (section X). | |
| Purified RNA is degraded. | RNases are present. | See “RNase elimination” in section VIII. Recommendations and Important Notes. |
| DNA contamination present. | Too much sample material was used. | Decrease the amount of sample material. Optionally, the purified RNA sample may be treated with a DNase. |
QUALITY CONTROL
The quality of each production batch (LOT) of the EXTRACTME VIRAL RNA KIT is tested with the use of standard QC procedures. Purified RNA concentration and quality are evaluated by gel electrophoresis and spectrophotometer
EXTRACTME VIRAL RNA KIT (EM39) - MSDS
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