Gel electrophoresis is a technique to separate and identify molecules. It has a wide variety of uses, from DNA analysis in microbiology labs to tests to identify drug use in pathology labs.
As one of Australia’s leading suppliers of gel electrophoresis equipment, we’ve put together a guide that will take you through the essentials of gel electrophoresis, explaining what it is and how it works. We’ll also elaborate on the many applications of gel electrophoresis to help you understand the application in a lab context.
What Is Gel Electrophoresis?
Gel electrophoresis is a laboratory technique used to separate mixtures of DNA, RNA, or proteins based on their size and charge. It allows the user to sort molecules, separating them to distinct categories.
The technique involves running an electric current through a gel matrix. This matrix acts like a sieve, allowing smaller molecules to move faster than their larger counterparts. The result is a separation, which enables researchers to analyse specific fragments of genetic material or proteins.
What Is Gel Electrophoresis Used For?
Let’s explore what gel electrophoresis is used for. This versatile technique has many uses in life science research.
1. DNA Analysis
One of the most common uses is DNA analysis. Researchers use gel electrophoresis to separate DNA fragments generated by restriction enzymes during cloning experiments. It’s a fundamental step in creating recombinant DNA, helping scientists identify and isolate specific genes.
2. RNA Analysis
In RNA research, gel electrophoresis helps assess RNA integrity. By analysing the ribosomal RNA bands, researchers can ensure the quality of their RNA samples, crucial for downstream applications like reverse transcription and quantitative PCR.
3. Protein Separation
For protein studies, gel electrophoresis plays a pivotal role in separating proteins by size, especially when combined with other techniques like Western blotting. This helps researchers study protein expression levels, modifications, and interactions.
4. Lab Diagnostics
In clinical settings, gel electrophoresis is a valuable method for diagnosing genetic disorders. Techniques like Southern blotting and Northern blotting rely on electrophoresis to detect specific DNA or RNA sequences associated with diseases.
5. Forensics
Gel electrophoresis is the backbone of DNA fingerprinting, helping match suspects to crime scene evidence with high precision.
6. Drugs Detection
Gel electrophoresis can identify narcotics present by analysing blood or urine samples.
7. Impurity Profiling of Pharmaceuticals
Yet another application of gel electrophoresis is to detect impurities in pharmaceutical products, ensuring their safety and fitness for use.
The Purpose of Gel Electrophoresis
We’ve touched upon its various applications, but what is the overarching purpose of gel electrophoresis? At its core, this technique serves as a vital method for analysing the composition, purity, and identity of nucleic acids and proteins.
By providing a visual representation of molecular separation, gel electrophoresis allows researchers to:
- Verify the success of molecular cloning experiments.
- Assess the size and quantity of nucleic acids or proteins.
- Identify mutations or variations in genetic material.
- Compare protein expression levels across different samples.
Its ability to provide insights into the molecular world makes it an essential technique in labs worldwide.
What equipment or supplies are used during gel electrophoresis?
A variety of equipment is required and the exact supplies will come down to your unique application. Here’s a broad overview of some of the different supplies involved in gel electrophoresis.
Electrophoresis Gel
A gel is used in electrophoresis to separate molecules, such as DNA, RNA, or proteins, based on their size and charge by applying an electric field that causes them to migrate through the gel matrix, allowing for analysis and identification. Common gels include:
- Agarose Gel (for DNA/RNA): The gel matrix used for separating nucleic acids. Agarose is a polysaccharide derived from seaweed, and it’s often dissolved in a buffer solution to form the gel.
- Polyacrylamide Gel (for proteins): Used for high-resolution separation of proteins and small nucleic acids. It is made by polymerising acrylamide and bisacrylamide in a specific ratio.
Electrophoresis Buffer Solutions
Typically, two main buffers are typically used: one for the running of the gel (running buffer) and one for preparing the sample (sample loading buffer)
- Running buffer: This buffer surrounds the gel during electrophoresis and facilitates the conduction of electricity. It also helps maintain the pH of the system to ensure that the macromolecules (DNA, RNA, or proteins) migrate correctly.
- Sample Loading Buffer: This buffer is mixed with the sample before loading it into the gel wells, serving to track migration with a loading dye, ensuring the sample sinks using a density agent like glycerol.
Electrophoresis tools
Several tools are needed to perform gel electrophoresis. These include but are not limited to:
- Gel Casting Tray: A flat tray or mold used to prepare and solidify the gel.
- Gel Electrophoresis Chamber: The chamber holds the gel and the buffer, and provides the electrical field necessary for electrophoresis. It connects to a power supply.
Power Supply: Provides current to move samples through the gel matrix. The voltage and current are adjustable depending on the sample type and the desired separation.
How Does Gel Electrophoresis Work?
Now that you’ve learnt a little more about gel electrophoresis, here’s an outline of how it works. To understand how gel electrophoresis works, let’s break down the various steps that make up the process.
1. Preparation of Samples
First, researchers prepare their samples, which might contain DNA, RNA, or proteins. The molecules are isolated and the samples are mixed with a loading dye, allowing them to be visualised as they migrate through the gel.
2. Gel Preparation
A gel is prepared, typically using agarose for DNA and RNA or polyacrylamide for proteins. The concentration of the gel can be adjusted based on the size of the molecules being analysed.
3. Casting the Gel
The gel is poured into a casting tray with wells moulded into it. Once it has cooled and solidified, this gel will serve as the medium through which the molecules will travel.
4. Loading the Samples
Prepared samples are carefully loaded into the wells. A molecular-weight size marker to enable identification of molecule sizes may also be added.
5. Running the Gel
The samples are placed into an electrophoresis chamber filled with buffers. The samples are positioned so that the wells are closest to the negative electrode. An electric current is applied across the gel. Molecules move towards the positive electrode, as they’re negatively charged thanks to the sugar-phosphate backbone of DNA and RNA or the inherent charge of proteins.
6. Visualisation
After the run is complete, the separated molecules are stained to make them visible. For DNA and RNA, stains like ethidium bromide or SYBR Green are used, while proteins might be stained with Coomassie Brilliant Blue or silver stain.
Tips for Successful Gel Electrophoresis
While the concept of gel electrophoresis is straightforward, achieving consistent results requires precision. Here are some tips to ensure success:
1. Optimise Gel Concentration
Choose the right gel concentration based on the size of the molecules you’re studying. Higher concentrations are ideal for smaller molecules, while larger molecules require lower concentrations.
2. Maintain Consistent Voltage
Running the gel at a consistent, optimal voltage ensures even migration of samples, preventing band smearing or distortion.
3. Use Fresh Reagents
Ensure all reagents, especially buffer solutions and staining agents, are fresh and properly prepared.
4. Handle Samples Carefully
When loading samples, avoid introducing bubbles into the wells. Bubbles can disrupt the migration pattern and lead to uneven bands.
5. Document Results Immediately
Once the run is complete and bands are visible, document results promptly with imaging equipment. Stains can degrade over time, affecting visibility.
Need to stock your lab for gel electrophoresis? Contact Fisher Biotec!
Gel electrophoresis is a workhorse in the lab, a trusted technique that continues to provide invaluable insights into the molecular realm. With a host of applications, it’s versatile too. So, whether you’re working with DNA, ensuring RNA quality, or studying protein interactions, consider using this technique for reliable results.
It’s also fast, simple, and low-cost, and analysis can be performed with only a small amount of sample. These benefits mean that gel electrophoresis is an excellent choice to get the rapid, reliable results you need with minimum effort.
Need lab supplies for gel electrophoresis? Fisher Biotec has you covered! We offer a comprehensive range of high-quality products, from agarose and precast gels to power supplies and electrophoresis chambers, ensuring you have everything needed for optimal performance. Contact us today and discover how we can help you enhance your gel electrophoresis workflow!