Extracting RNA from diverse sample types presents significant challenges that can impact downstream applications. Regardless of the method used, several key factors must be addressed to ensure high-quality, intact RNA:
RNA Integrity & Stability: RNA is highly susceptible to degradation, so preserving its integrity is critical during extraction. To avoid RNA integrity loss, it is essential to apply a rapid processing workflow, minimise exposure to RNases enzymes and maintain optimal low-temperature conditions throughout the process.
Contamination Risks: DNA, proteins, and other impurities can interfere with RNA purity and consequently downstream applications. Effective purification techniques are crucial to eliminate these contaminants.
Optimising Yield: Certain sample types — such as blood, bacteria, or tough tissues — yield low RNA amounts, requiring optimised protocols, larger sample volumes, or concentration techniques.
Extracting RNA from infectious disease samples adds another layer of complexity, requiring effective pathogen inactivation to ensure safe handling of potentially contaminated biological samples with bacteria, viruses, and yeast.
Addressing these challenges ensures the extraction of high-quality, contamination-free RNA, leading to reliable and reproducible results in molecular studies.
RNA Extraction using TRIzolâ„¢
TRIzolâ„¢ is a monophasic solution containing phenol, guanidine isothiocyanate, and proprietary components that facilitate the isolation of RNA species of various sizes. The solution also contains sodium acetate and has a pH of 3. The name itself is derived from the method of isolating RNA, DNA, and proteins in three separate phases. TRIzolâ„¢ can be used with various sample types, including cells, solid tissues, blood, plants, and infectious biological samples.
TRIzolâ„¢ is particularly well-regarded for RNA extraction from infectious biological samples. There are several reasons for this:
1. Viral and Bacterial Inactivation
TRIzol™ is highly effective at inactivating infectious agents, including viruses and bacteria. A 2023 study published in Applied Biosafety by Gwen Duytschaever et al. demonstrated TRIzol™’s effectiveness in neutralizing pathogens, including Ebola. The study concludes:
Available evidence suggests that TRIzol™ Reagent is effective in inactivating a broad spectrum of bacteria and viruses from cells, tissues, and liquids in biological samples when the matrices are exposed to at least 10 min at room temperature to the reagent…
2. RNA Stability in TRIzolâ„¢
TRIzolâ„¢ stabilizes RNA by inactivating RNase enzymes through its unique combination of guanidine isothiocyanate, phenol, and acidic pH (3), eliminating a key cause of RNA degradation. This ensures that RNA remains intact and suitable for downstream applications.
By leveraging these properties, TRIzolâ„¢ has become a trusted reagent for RNA extraction, particularly in handling infectious disease samples where both pathogen inactivation and RNA integrity are critical.
Microzone’s MyZol for TRIzol™
MAGneat Bead RNA extraction from TRIzolâ„¢ reagent
While TRIzolâ„¢ offers numerous benefits for RNA extraction, researchers often encounter challenges that impact its ease of use, efficiency, and effectiveness. This section highlights common difficulties associated with TRIzolâ„¢ and explains how MyZol for TRIzolâ„¢ MAGneat RNA Extraction Kit helps overcome these obstacles.
1. Incomplete Phase Separation
After chloroform addition, incomplete separation of aqueous and organic phases can lead to RNA contamination with DNA or proteins, reducing RNA yield.
MyZol eliminates the need for phase separation, preventing contamination and improving RNA purity.
2. Low RNA Yield
Poor RNA recovery can result from incomplete cell lysis — especially in hard-to-lyse tissues or bacteria — or poor RNA solubility.
MyZol consistently yields higher RNA recovery compared to TRIzolâ„¢ and alternative bead-based methods.
Table 1. MyZol for TRIzol — MAGneat RNA Extraction Kit outperforms other commercially available TRIzol RNA extraction kits. RNA was extracted from avian muscle tissue in TRIzol, using the MyZol for TRIzol — MAGneat RNA Extraction Kit and a commercially available comparator.
3. Contamination with DNA or Protein
RNA extracted with TRIzolâ„¢ may contain DNA or proteins, affecting downstream applications like qPCR or Northern blotting.
MyZol is bead-based and selectively captures only nucleic acids. With the addition of a DNase step MyZol ensures that the final product is free of both DNA and protein and RNA rich.
4. Inconsistent Results
Variations in sample type, sample size, or reagent conditions can lead to inconsistent RNA yield or quality.
MyZol provides reliable results in both yield and purity, with the added advantage of automation for enhanced reproducibility.
5. Chloroform Fumes
Chloroform, a hazardous chemical used in TRIzolâ„¢ extraction, produces toxic fumes that pose health risks.
MyZol is a chloroform-free system, offering a safer and more user-friendly alternative.
6. Trouble with Precipitation Step
RNA precipitation with isopropanol can be inefficient due to improper ethanol washing or excessive sample volume.
MyZol eliminates the need for a precipitation step, simplifying the workflow and improving consistency.
7. Requirement for Low-Temperature Centrifugation
TRIzolâ„¢ protocols recommend centrifugation at 4ËšC to facilitate phase separation.
MyZol eliminates the need for expensive refrigerated centrifuges, reducing equipment dependency.
8. Speed and Automation
MyZol is a rapid and scalable method, suitable for manual use in small-scale experiments or automation in high-throughput systems like the HeiDi-NA (1–16 samples) or KingFisher (96 samples at a time).
Conclusion
TRIzolâ„¢ is a powerful and reliable reagent for RNA extraction, especially valuable when working with infectious biological samples due to its ability to inactivate pathogens. Its effectiveness in protecting RNA integrity by neutralizing RNases has made it a go-to choice for researchers. However, traditional TRIzolâ„¢ protocols can present challenges such as incomplete phase separation, low RNA yield, contamination with DNA or protein, and the use of hazardous chemicals like chloroform.
MyZol for TRIzolâ„¢ MAGneat RNA Extraction Kit offers a streamlined and safer alternative, enhancing RNA yield and purity while eliminating common extraction issues. Its ease of use, higher reproducibility, and automation potential make it an excellent choice for researchers looking to optimise their RNA extraction workflow, whether for small-scale or high-throughput applications.
Ready to enhance your RNA extraction process? Contact us today via the contact form to learn more and discuss how MyZol can help you achieve more consistent, efficient results.
Product Spotlight
MyZol for TRIzol – MAGneat RNA Extraction Kit
The MyZol Kit provides efficient RNA extraction from samples stored in TRIzol, including cells, viruses, tissues, and biological liquids. Utilizing MAGneat magnetic bead technology, the kit enables selective and reversible RNA binding, achieving superior yields compared to spin-column methods. Microzone’s proprietary buffers ensure the removal of contaminants, delivering high-purity RNA suitable for sensitive downstream applications.
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