Question

In phage display using a phagemid vector derived from M13 filamentous phage, you need to transfect a helper phage to propagate the phagemid vector. Explain why ?

Answer 1

Phage display is a powerful technique to know about DNA protein or protein protein interaction with huge industrial application. A phagemid is used during such screening. Phagemid is a plasmid with one extra extra origin of replication for the phage apart from usual origin of replication site for the plasmid. The phagemid has property of both bacteriophage as well as of plasmid

During phage display - sequence for desired gene is fused with gene of coat protein of the phage (pIII or PVIII in case of M13 phage).

So, that after protein synthesis the product of our gene of interest will have a fused coat protein as well

Fused coat protein will cause our protein of interest to be inserted on coat of phage (hence the name phage display)

Now, when we insert our gene of interest in phagemid. The phagemid contains only two things mainly viral coat protein and our desired gene/protein.

Afterward, we insert this phagemid in Bacterial host E.coli by transduction.

Now, since the phagemid has only two genes -coat protein (pIII or PVIII in case of M13 phage) and Gene Of Interest along with two origin of replication. It will not be enough information for packaging, assembly and release of phage particles from the bacterial host. To help phagemid in performing these processes, we infect the bacteria with a helper phage. Helper phage helps the deficient plasmid in assembly, packaging and release of new virions. Therefore, infection with helper phage is vital. If helper phage will not be used then packaging of Phagemid and release of complete virion will not be possible.

After mature virions are released we screen our protein of interest for its ability to bind on respective site or surface (covered with specific DNA or protein). Then we screen and elute the bound phage and We can increase their number by again infecting a bacterial host.