While DNA can remain stable for many years when stored under appropriate conditions, RNA is short-lived. Because enzymes called “RNase”, which can be found in all kinds of conditions, cause RNA degradation. Therefore, a method that can isolate total RNA from cells with high efficiency and at the same time prevent RNA degradation is required.
What is total RNA?
Total RNA is all the RNA molecules present inside a cell. All prokaryotic and eukaryotic cells contain the following types:
Messenger RNA (mRNA): A long protein-coding messenger RNA that carries to DNA and serves as an instant readout of cellular gene expression under certain conditions.
MicroRNA (miRNA): Countless other non-coding RNA molecules, many of which are involved in the regulation and silencing of gene expression.
Ribosomal RNA (rRNA): An important component of ribosomes and critical for protein synthesis.
Transfer RNA (tRNA): Another critical component for protein synthesis. It is formed by processing a precursor molecule in the nucleus. These RNA molecules carry amino acids to the ribosome and base pair with mRNA to ensure that the correct amino acid is added to the synthesized protein.
RNaz What is it?
Ribonucleases (RNases) are enzymes that cleave RNA. These enzymes are very problematic during RNA isolation because they are ubiquitous and very difficult to eliminate. When isolating total RNA you need to make sure that you eliminate the RNAase.
Total RNA Isolation RNases Trizol Removing Using
Inactivating RNases is, of course, not impossible. The first method to safely isolate RNAs while inhibiting RNases was the Guanidinium thiocyanate-phenol-chloroform isolation method applied by PiotrChomczynski and Nicoletta Sacchi. This method is a form of a potent protein denaturing chemical that breaks down protein cell components and inactivates all enzymes including RNases. Isolation typically uses acidic phenol-chloroform to confine total RNA in a clear aqueous phase, while proteins and cell debris end up in the pink organic layer. The RNA can be recovered by precipitation with isopropanol, washed and then redissolved in water. There are many ready-made brands of this method. And it is also possible to make your own phenol guanidine isothiocyanate mixture.
For total RNA isolation Trizol’s Benefits
Denaturing RNases.
Isolation of total RNA, including small molecular weight RNAs such as miRNA
High quality RNA.
It is relatively simple to use.
It allows simultaneous isolation of DNA, protein and RNA from a sample.
Trizol Disadvantages of Using
It can be costly compared to other methods of RNA isolation,
Dangerous and toxic,
If not handled carefully, isolated RNA may be contaminated with phenol during phase separation,
It can be time-consuming.
For RNA isolation Trizole Alternatik Commercial Kits Used as
There are many commercial kits that do not use trizol or other hazardous chemicals. Today, commercial kits are available that are suitable for almost all applications. These kits can use spin column or magnetic bead methods to capture RNA. There are also kits specifically designed for miRNA or other small molecular weight RNA species. Kit methods are often safer and more stable than Trizol because they are optimized to extract RNA from different sources and materials.
SUMMARY
Trizol and commercially available kit methods have pros and cons in terms of cost, use of hazardous chemicals and convenience, each of which should be carefully considered before selection.