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product rather than the DNA inserted into the vector. This screening is therefore

undertaken with antibody probes directed against the protein of interest (Section

6.5.4). Other features that make this a useful cloning vector are the ability to produce

RNA transcripts termed cRNA orriboprobes. This is possible because two promoters

for RNA polymerase enzymes exist in the vector, a T7 and a T3 promoter which flank

the MCS (Section 6.4.2).

One of the most useful features oflZapisthatithasbeendesignedtoallow

automatic excisionin vivoof a small 2.9 kb colony-producing vector termed a phage-

mid, pBluescript SK (Section 6.3.3). This technique is sometimes termedsingle-stranded

DNA rescueand occurs as the result of a process termedsuperinfectionwhere helper

phage are added to the cells which are grown for an additional period of approximately

4 h (Fig. 6.19).

T I

DNA inserted into multiple cloning site

Phagemid

vector site

InfectE.coli

Add helper phage (e.g. M13R408)

Excision of phagemid from λZAP vector

f1(+/–) origin

Ampicillin

resistance

gene

Col E1

plasmid origin

DNA inserted into

Bluescript SK+/–2.96 kb multiple cloning site

Cos

Cos

A–J CI857

ApR lacZ

Fig. 6.19Single-stranded DNA rescue of phagemid fromlZap. The single-stranded phagemid pBluescript SK may be

excised fromlZap by addition of helper phage. This provides the necessary proteins and factors for transcription

between the I and T sites in the parent phage to produce the phagemid with the DNA cloned into the parent vector.

215 6.3 Cloning vectors