Introduction
Northern blotting, a technique for analysis of RNA, was proposed by James Alwine, George Stark and David Kemp in 1977 at Stanford University. It is a method used for the detection of gene expression, specific RNA sequences and various RNA functions. It involves the movement of those RNA molecules on a solid matrix which were already separated by electrophoresis. The matrix is exposed to the suitable probe that hybridizes with the RNA sample. The detector (e.g. phosphoimager and X-ray) then detects the probe. The technique is named as Northern blotting in contrast to pre-existing method, Southern blotting, used for the DNA analysis.
The RNA molecule
RNA, a nucleic acid molecule, is made up of ribose sugar, phosphate backbone nitrogenous…show more content… 2nd cistron translation
• investigation of abnormal gene and genetic disorders
• plays a vital role in the study of certain proteins that are involved in cancerous growth
• normal and cancerous tissues can be compared
• study of oncogene overexpression and tumor gene suppression
Pros and Cons
The advantages and limitation of northern blotting are given below:
Pros
• It is well regarded and broadly accepted method.
• It is versatile as it permits the usage of different types of probes including: radiolabeled and non- radiolabeled, and even oligonucleotides like primers.
• It is a flexible and simple technique.
Cons
• Time consuming
• RNA samples are if degraded somewhat by RNases, the quality and quantity of expression is badly affected.
• Other specialized techniques such as nuclease protection assays and RT-PCR are required for absolute RNA quantification and high sensitivity as northern blotting technique is less sensitive. Its sensitivity can be enhanced by using nylon positively-charged membranes or extremely specific antisense