Nucleic Acids Research, 1988, Vol. 16, No. 15 7583-7600
© 1988
Articles |
ZAP: a bacteriophage expression vector with in vivo excision properties
Stratagene Cloning Systems 11099 North Torrey Pines Road, La Jolla, CA 92037, USA
*To whom correspondence should be addressed
Received February 16, 1988. Revised June 29, 1988. Accepted June 29, 1988.
A lambda Insertion type cDNA cloning vector, Lambda ZAP, has been constructed. In E. coli a phagemid, pBluescript SK(–), contained within the vector, can be excised by 11 or M13 helper phage. The excision process elimlnates the need to subclone DNA inserts from the lambda phage into a plasmid by restriction digestion and ligation. This is possible because Lambda ZAP incorporates the signals for both initiation and termination of DNA synthesis from the 11 bacteriophage origin of replication (1). Six of 21 restriction sites In the excised pBluescript SK polylinker, contained within the NH2-portion of the lacZ gene, are unique in lambda ZAP. Coding sequences inserted into these restriction sites, in the appropriate reading frame, can be expressed from the lacZ promoter as fusion proteins. The features of this vector significantly increase the rate at which clones can be isolated and analyzed.
The lambda ZAP vector was tested by the preparation of a chicken liver cDNA Hbrary and the isolatlon of actin clones by screening with oligonucleotide probes. Putative actin clones were excised from the lambda vector and Identified by DNA sequencing. The ability of lambda ZAP to serve as a vector for the construction of cDNA expression libraries was determined by detecting fusion proteins from clones containing glucocerbrosidase cDNA's using rabbit IgG anti-glucocerbrosidase antibodies.
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