Total Nucleic Acid Extraction Kit (Virus)
No:DMNAV100 & 300
For research use only
Sample: serum, plasma, body fluids and the supernatant of viral infected cell cultures
Introduction
The BioDiamond Virus Total Nucleic Acid Extraction Kit provides a fast, simple, and cost-effective method for isolation of viral DNA/RNA from cell-free samples such as serum, plasma, body fluids and the supernatant of viral infected cell cultures. Its unique buffer system efficiently lyses cells and degrades proteins, allowing for nucleic acids to bind to the glass fiber matrix of the columns easily. Contaminants such as salts, metabolites and soluble macromolecular cellular components are removed through the Wash step. Phenol extraction and ethanol precipitation are not required, and high-quality Nucleic Acids are eluted with RNase-free elution buffer. Viral DNA/RNA isolated with BioDiamond’s Virus Total Nucleic Acid Extraction Kits is suitable for a variety of routine applications, including Real-time PCR/RT-PCR, Automated Fluorescent DNA Sequencing, PCR, and other enzymatic reactions. The entireprocedure can be completed within 15-20 minutes.
Kit Contents
NOTE
★ Add ethanol (96–100%) to Buffer V2 and W2, shake before use (see bottle label for volume).
★ Check Buffers before use for salt precipitation. Redissolve any precipitate by warming to 37°C.
★ Buffers V1 and W1 contain irritants. Wear gloves when handling these buffers.
Quality Control
In accordance with FairBiotech’s ISO-certified Total Quality Management System, the quality of the BioDiamond Virus Total Nucleic Acid Extraction Kit is tested on a lot-to-lot basis to ensure consistent product quality.
Additional requirements
*absolute EtOH*PBS (Phosphate Buffered Saline)
*microcentrifuge tubes (DNase and RNase free)
Protocol
Step 1 Lysis
- Transfer up to 200 μl of virus sample into a 1.5 ml microcentrifuge tube and add 400 μl of V1 Buffer. (If the sample is less than 200 μl, adjust the sample volumn to 200 μl with PBS)
- Mix well and let stand at room temperature for 10 minutes.
Step 2 Nucleic Acid Binding
- Add 450 µl of V2 Buffer (ethanol added) to the sample lysate and shake vigorously.
- Place a VN Column in a Collection Tube. Transfer 700 µl of the lysate mixture to the VN Column.
- Centrifuge at 16,000 x g for 1 minute. Discard the flow-through and place the VN Column back in the Collection Tube.
- Transfer the remaining lysate mixture to the VN Column.
- Centrifuge at 16,000 x g for 1 minute. Discard the flow-through and place the VN Column back in the Collection Tube.
- Add 400 µl of W1 Buffer into the VN Column. Centrifuge at 16,000 x g for 30 seconds. Discard the flow-through and place the VN column back into the Collection tube.
- Add 600 µl of W2 Buffer (ethanol added) into the VN Column. Centrifuge at 16,000 x g for 30 seconds. Discard the flow-through and place the VN column back in the Collection tube.
- Centrifuge at 16,000 x g again for 2 minutes to remove residual W2 Buffer .
- Place the VN column in a clean 1.5 ml microcentrifuge tube (DNase and RNase free).
- Add 50-100 µl of Pre-Heated EL Buffer or RNase-free water (pH is between 7.0 and 8.5) to the center of each VN column, let stand for 2 min, and centrifuge at 14,000 x g for 2 min.
Troubleshooting
dmnav100___300.pdf | |
File Size: | 243 kb |
File Type: |