Difference between southern bloting and nortjern bloting
Answers
What is Southern Blotting?
Southern blotting technique was developed by E. M. Southern in 1975 for the identification of a specific DNA sequence from a DNA sample. This is the first blotting technique introduced in molecular biology. It enabled the detection of specific genes from the DNA, specific fragments from DNA, etc. There are several steps involved in southern blotting technique. They are as follows.
DNA is isolated from the sample and digested with restriction endonucleases.
Digested sample is separated by Agarose gel electrophoresis.
DNA fragments in the gel are denatured into single strands by the use of an alkaline solution.
Single stranded DNA is transferred to a nitrocellulose filter membrane by capillary transferring.
Transferred DNA is fixed onto the membrane permanently.
Fixed DNA on the membrane is hybridized with labeled probes.
Unbound DNA is washed off from the membrane by washing.
X-ray film is exposed to the membrane and an autoradiograph is prepared
What is Northern Blotting?
Northern blotting is a method designed to detect a specific RNA sequence or mRNA sequence from a sample to study gene expression. This technique was developed by Alwine, Kemp, and Stark in 1979. It differs from the sourthern and western blotting techniques due to several steps. However, this technique is also performed via gel electrophoresis, blotting, and hybridization with specific labeled probes and detection. Northern blotting technique is performed as follows.
RNA is extracted from the sample and separated by gel electrophoresis.
RNA is transferred from the gel onto a blotting membrane and fixed.
The membrane is treated with a labeled probe prepared from cDNA or RNA ( the probe is complementary to a specific sequence in the sample).
The probe is incubated with the membrane to bind with a specific sequence.
Unbound probes are washed off.
Hybridized fragments are detected by an autoradiograph.
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