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DNA/RNA Shearing Protocols
DNA/ RNA shearing protocols for the various Focused-ultrasonicator instruments and Covaris certified consumables.
AFA Process for DNA and RNA Shearing
Adaptive Focused Acoustics® (AFA®) technology is firmly established as the fragmentation method of choice for NGS, and Covaris continues to innovate the tools required to shear DNA and RNA without GC bias or thermal damage.
The Covaris AFA process is conducted under isothermal conditions, ensuring the integrity of the nucleic acid sample is maintained and providing high recovery of double-stranded DNA during the shearing process.
Combined with the specifically engineered AFA Tubes it is possible to precisely and accurately fragment DNA and RNA to the 100 – 1500bp range (microTUBE), or 2 – 5kb range (miniTUBE). Focused-ultrasonicators range from the low through-put single-tube full-power S-Series, the mid-throughput, more affordable M-Series, to the higher throughput E and L-Series instruments.
Benefits of RNA or DNA Shearing using AFA Technology
Physical shearing of DNA or RNA using Covaris AFA is concentration independent, isothermal, and highly reproducible. Physical shearing does not require a cleanup step after shearing, thereby preserving the complexity of the sample for library preparation. This shearing method provides unmatched control and reproducibility of shearing.
| FEATURE | BENEFIT |
| Temperature Controlled | Processing at controlled temperatures provides highest yields while preserving sample fidelity |
| Accurate and precise | Highly reproducible process, day to day, user to user. Protocols can be transferred instrument to instrument without further optimization |
| Non-contact, closed vessel | No cross-contamination, clean-up, or sample loss |
| Versatile | Generate tight fragment distribution centered from 100 bp to 20 kbp |
DNA Shearing of Clinical Samples
Unparalleled acoustic energy control enables shearing across a wide range of fragment sizes ranging from 150 bp to 5 kb and everything in between. AFA-energetics is the gold standard for DNA shearing providing you with highest sample complexity and unbiased sample yield paving the way for the most cost-effective library prep.
RNA Shearing
Total RNA sheared with Covaris AFA from FFPE samples. These electropherograms show unbiased RNA fragment size distribution.
Total mRNA Shearing
Agilent BioAnalyzer electropherogram of 5 µg of mRNA processed in a volume of 130 µl of TE in a microTUBE according to the operating conditions, showing the mean fragment size at 200 bases.
Comparison of Shearing Fragment Sizes
| Fragment Size* | Sample Volume | Covaris Products |
| 175 to 1.5 kbp | 5 to 50 µl | AFA-TUBE |
| 150 to 1.5 kbp | 130 µl | microTUBE-130 |
| 150 to 1.5 kbp | 55 µl | microTUBE-50 |
| 150 to 1.5 kbp | 15 µl | microTUBE-15 |
| 2 kbp | 200 µl | miniTUBE-Clear |
| 3 kbp | 200 µl | miniTUBE-Blue |
| 5 kbp | 200 µl | miniTUBE-Red |
| 6 – 20 kb | 150 µl | g-TUBE |
*For validated fragment sizes, refer to the Covaris Focused-ultrasonicator Protocol.
Featured Products
AFA-TUBE
8 strip and plate-based engineered polymer vessels with RFID enabled sample tracking.
microTUBE
Glass tubes specifically engineered for use with Focused-ulrasonicators. Meant for small sample volumes, 130µl or less.
