Phage vectors, cosmids and phasmids

Viruses that infect bacteria, bacteriophages, make good vectors too. They can carry larger inserts than plasmids, and their plaques (clear spots where they have killed bacteria on a plate) are easier to screen than bacterial colonies. The two most used, both for E. coli, are lambda and M13.

Phage lambda has a head, holding its DNA, and a tail. It can follow either of two life cycles: lytic (it multiplies and bursts the cell) or lysogenic (its DNA joins the host chromosome as a prophage and waits). Its DNA is linear, 48,490 bp long, with 12-base single-stranded ends, the cos sites, which stick together to close it into a circle inside the cell.

About one third of the lambda genome, the middle region, holds genes for the lysogenic cycle, which are not needed to grow the phage in the lytic cycle. So this part can be replaced by an insert. But the total DNA must stay between 38 and 52 kbp, or it will not fit into the phage head.

• Insertion vectors have one cloning site, into which the insert is placed.
• Replacement vectors have a removable stuffer region, which is swapped for the insert.

Phage M13 is a thread-like (filamentous) phage with a single-stranded, circular genome of 6407 bp. It infects only E. coli cells that carry the F plasmid, entering through their pilus. Inside, it makes a double-stranded replicative form, which is used as the vector; the insert goes into the region between its genes. Unusually, M13 leaves the cell without bursting it. M13mp18 is designed to allow blue/white selection.

Two hybrid vectors borrow from both:

• A cosmid is a plasmid that carries lambda's cos site. It replicates like a plasmid, but can be packed in vitro into lambda coats, which deliver it efficiently. It takes inserts of up to about 45 kbp.
• A phasmid has lambda's two ends, with all its lytic genes, and a linearised plasmid in the middle. It can grow like a phage, or like a plasmid if it carries the lambda repressor gene.