Northern Blot- Definition, Principle, Steps, Results, Applications


RNA is a type of nucleic acid that carries the genetic information from DNA to the protein synthesis machinery in the cell. RNA molecules can have different functions and structures, such as messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), and microRNA (miRNA). The expression of RNA can vary depending on the cell type, tissue, developmental stage, and environmental conditions. Therefore, studying RNA can provide insights into the regulation of gene expression and the molecular mechanisms of various biological processes and diseases.

However, analyzing RNA can be challenging because RNA molecules are often present in low amounts and are easily degraded by enzymes called RNases. Moreover, RNA molecules can have different lengths and sequences, which makes it difficult to identify and separate them from a complex mixture. To overcome these challenges, molecular biologists have developed a technique called Northern blot, which allows the detection and quantification of specific RNA molecules among a mixture of RNA.

Northern blot is named after its similarity to another technique called Southern blot, which is used to analyze DNA. Northern blot involves four main steps:

  1. Separation of RNA on a denaturing gel by electrophoresis
  1. Transfer of RNA from gel to a nylon membrane by capillary action
  1. Immobilization of RNA on the membrane by UV cross-linking or baking
  1. Hybridization of the membrane with a labeled probe that is complementary to the target RNA sequence

The labeled probe can be detected by various methods, such as autoradiography, chemiluminescence, or fluorescence. The intensity and position of the signal on the membrane indicate the amount and size of the target RNA molecule in the sample.

Northern blot is a powerful technique that can be used for various purposes, such as:

  • Identifying and characterizing different types of RNA molecules, such as mRNA, tRNA, rRNA, and miRNA
  • Measuring the expression levels of specific genes in different tissues or cells
  • Comparing the expression patterns of genes under different conditions or treatments
  • Detecting mutations, deletions, insertions, or splicing variants in RNA sequences
  • Studying the structure and function of RNA molecules

In this article, we will explain the principle, equipment, materials, steps, results, applications, and limitations of Northern blot in detail. We will also provide some examples and tips for performing this technique successfully.