Effective separation and detection of RNA and DNA in molecular biology research is essential for gene discovery, sequencing, and mapping used in diagnostics and industrial applications. Northern and Southern blotting are standard molecular biology techniques for RNA and DNA identification and quantification, respectively.
Alfa Chemistry strives to simplify detection through consistent, easy-to-read Northern and Southern blot transfer. We provide highly sensitive transfer membranes for Northern and Southern blot detection methods. It can identify and quantify RNA and DNA even in low-abundance samples.
Clarity of Visual Results
Northern blotting is used to detect specific RNA sequences in a variety of RNA mixtures, which involves complex separation and hybridization procedures that result in the binding of labeled probes to the RNA sequence of interest for detection and visualization. Southern blotting is used to detect specific DNA sequences in large and complex DNA samples. It can also be used to determine the molecular weight of restriction fragments and measure the relative amount of DNA in different samples.
Figure.1 Steps involved in DNA/RNA blotting procedure
Northern and Southern blotting analysis methods are based on the immobilization of nucleic acids. RNA and fragments are separated by size on the membrane to allow the use of macromolecular probes for identification, detection, and visualization. To obtain clear results, the transfer film used needs to have:
- High sensitivity
- Surface consistency
- Correct sensitivity level
- Low background noise
When the abundance of RNA and DNA molecules is low, they may be difficult to separate and visualize. Membrane and detection schemes must provide the sensitivity to detect small amounts of molecules of interest without background interference. Alfa Chemistry provides transfer membranes with high sensitivity, low background and batch-to-batch consistency for all radioactive and non-radioactive detection methods.
The Right Membrane for Your Application
The nylon transfer membrane series provides a series of chemical substances with a variety of adsorption characteristics, suitable for different applications. These membranes are hydrophilic in nature, so it is easy to wet the entire membrane. Their stability and durability ensure that they will not rupture, shrink, or tear during multiple cycles of hybridization, peeling, and reprobing.
A membrane zeta potential can be adjusted by pH value, which is very suitable for a single probe or multiple rehybridization. Membrane B is a positively charged nylon membrane with the highest sensitivity for nucleic acid applications. Plus membrane has a very high isoelectric point, which means it will maintain a good charge as the pH changes. C membrane is suitable for professional applications because it can be derivatized by the coupling reaction of carboxyl groups on the pore surface.
Please refer to the following selection guide for more information.
Northern and Southern Transfer Membrane Selection Guide | |||
Membrane A | Membrane B/Plus | Membrane C | |
Description | Amphoteric Nylon 6,6 | Positively-charged Nylon 6,6 | Negatively-charged Nylon 6,6 |
Advantages | High sensitivity Low background Ability to strip and reprobe Net change can be controlled by changing pH | Positive charge over broad pH range Ability to strip and reprobe Highest sensitivity for nucleic acid applications (membrane B) | Ability to strip and reprobe Negative charge over broad pH range Surface carboxyl groups can be derivatized |
Works suited for | Colony/Plaque Lifts DNA and RNA Transfers Gene Probe Assays, DNA Fingerprinting, Nucleic acid Dot/Slot Blots, Replica Plating, ELISAs | Positive charge over broad pH range Ability to strip and reprobe Highest sensitivity for nucleic acid applications (membrane B) | Reverse Dot Blots Protein Immobilization Affinity Purification ELISAs |
Detection Methods | Radiolabeled Probes, Enzyme-antibody Conjugates
| Radiolabeled Probes, Enzyme-antibody Conjugates
| Radiolabeled Probes, Enzyme-antibody Conjugates
|
Binding Interaction | Hydrophobic and Electrostatic | Hydrophobic and Electrostatic | Hydrophobic and Electrostatic |