Northern blotting is a technique used to detect specific mRNAs in a sample. It involves separating RNA fragments by size through gel electrophoresis, transferring them to a membrane, then using a complementary DNA or RNA probe to identify the mRNA of interest. It allows researchers to examine gene expression by measuring mRNA levels and determine tissue specificity and regulatory factors of gene expression. While it provides useful information, it also has some limitations such as risk of mRNA degradation and relatively low sensitivity compared to PCR-based methods.

1. NORTHERN
BLOTTING
TECHNIQUE
Prepared By...
Nabajit Roy
M.Sc 2nd Semester
Paper –LS-202
Dept. of Life Science and Bioinformatics
AUS
2017-2018

2. CONTENTS
 What is blotting
 Types of blotting
 Principle
 Northern blotting
 Application
 Advantage & Disadvantage
 Summary
 References
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3. What is Blotting?
 In molecular biology blotting is a technique for transferring DNA , RNA
and proteins onto a carrier so they can be separated, and often follow
the use of gel electrophoresis.
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4. Types OF Blotting:
Types of blotting techniques are differentiated by the presence of the target molecules
that is being sought. The first of this techniques developed was Dr. Edwin Southern
(1975).
 Southern Blotting: Used For DNA Detection in a sample
 Northern Blotting : Used For RNA Detection in a sample
 Western Blotting : Used For Proteins in a sample
 Dot blotting technique: to detect the presence of a given sequence of
DNA/RNA
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5. PRINCIPLE
1. RNA are separated by size and detected on a membrane
using a hybridization probe with a base sequence
complementary to all, or a part, of the sequence of the
target mRNA
2. transfer of RNA from a gel to a membrane.
3. And then the membrane is queried with a probe directed
against the specific molecule of interest.
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6. NORTHERN BLOTTING
 The Northern blot is a technique used in molecular biology to study gene
expression by detection of RNA (or isolated mRNA) in a sample
developed by Alwnie and his colleagues in 1979. This method was named
for its similarity to the technique known as a Southern blot.
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8. APPLICATION
 Detecting a specific mRNA in a sample
 Used in the screening of recombinants by detecting the mRNA produced
by the transgene.
 In disease diagnosis.
 In gene expression studies.
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9. ADVANTAGE
1. The strength of this technique is its
simplicity.
2. Specificity is relatively high.
3. mRNA transcript size can be detected.
4. The cost of running many Gels is low
once the equipment is setup.
5. Blots can be stored for several years and
reprobed if necessary.
6. Quantity and Quality of RNA can be
easily verified after electrophoresis and
before further processing is done.
DISADVANTAGE
1. Risk of mRNA degradation during
electrophoresis: Quality &
Quantification of expression are
negatively effected.
2. High doses of radioactivity and
formaldehyde are a risk for workers and
the environment.
3. The sensitivity of Northern blotting is
relatively low in comparison with that
of RT-PCR.
4. Detection with multiple probe is
difficult.
5. Use of Ethidium Bromide (EtBr), DEPC
& UV light needs special training and
attention
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10. SUMMARY
 Northern blotting is the main method for examining the expression of genes through
measurement of an mRNA. Or It is theoretically a good technique for determining the no. of
genes (through mRNA) present on a given DNA. Tissue specificity and the factors which
regulate expression can both be determined by Northern blotting. A cDNA radioactively
labelled with 32P is the most-commonly-used hybridization probe. Convenient, non-
radioactive detection protocols are, however, increasingly available. A combination of anti-
sense oligonucleotides as probes, together with chemo-luminescence-based detection
provides a rapid and simplified approach to Northern blotting, increasing the accessibility of
this important procedure for nutritional studies.
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11. REFERENCES
1. Wilson, K & Walker, J,(2010), Principles and techniques of biochemistry and molecular
biology.
2. Satyanarayan, U.(2015) biotechnology pp. 68-101
3. Goda, S.K. & Nigel, P.M., “A simple procedure for gel electrophoresis and northern
blotting of RNA”(1995) “Nucleic acid research 23.16:3357
4. Trayhurn, P.(1996). Northern Blotting Proceedings of the Nutrition society. Pp 55, 583-589.
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