Question

What is Inverse PCR?

Answer 1

Hint: Polymerase Chain Reaction (PCR) is a laboratory technique in which DNA fragments are amplified using complimentary polymers specific to DNA sequences in enzymatic cyclic reaction, which is temperature dependent. Inverse PCR is just a variant of conventional PCR. Inverse PCR was developed by Howard Ochman and Coworker in \[1988\].

Complete answer:
Target DNA is known to us in the conventional PCR. But in inverse PCR, the target DNA is unknown to us.
There are five steps involved in the inverse PCR process.
Identification of target sequence flanked by unknown DNA sequence
Restriction digestion of genomic DNA.
Ligation of unknown DNA sequences
Amplification of unknown DNA sequences through known DNA regions.
Sequencing of unknown DNA sequences.
Inverse PCR involves digestion and self-ligations of DNA cut by restriction endonuclease. Cut gives rise to known sequences at both ends of unknown sequences. It differs from the conventional PCR in that the primers are oriented in the reverse direction. Template for the primers is a restriction fragment ligated upon itself forming a circle.
The brief procedure is as follows:
Target DNA is cut into smaller fragments. Under lower concentrations, self-ligation is induced, causing formation of phosphate backbone. It leads to the formation of circular DNA products. This is cut using known restriction nucleases. A cut is generated within the known DNA sequence, leading to formation of linear products with known sequences. Then, this product is used for conventional PCR reactions.

Note:
Inverse PCR is used to find out the flanking sequences, which are present around genomic inserts. It is helpful in identification and amplification of sequences, identification of genomic inserts and flanking transposable elements. It also helps in investigation and identification of DNA downstream and upstream region to the exon. Besides, it also helps to identify unknown gene mutations like gene fusion, gene rearrangement and oncogenic gene arrangement.