Experimental Methods for Use with Lambda

Abstract

INTRODUCTION
This paper is addressed particularly (but not exclusively) to those who use λ as a vector for cloning genes of other organisms. In it we describe techniques for growing and purifying phage and offer a brief menu of tests that are useful for verification of genetic characters found in some widely used vectors. We also suggest methods for simple strain construction, mutagenesis, DNA purification, and in vitro DNA packaging. Alternative and overlapping procedures can also be found (Miller 1972; Davis et al. 1980; Maniatis et al. 1982; Berman et al. 1983). We emphasize that the choice of route is often a matter of individual preference and that methods can and should be varied to suit the particular phage strain, laboratory conditions, and the convenience of the investigator. We have indicated why certain manipulations are to be preferred or avoided, but we warn the reader that the reasons are often speculative and are presented to demystify our subject rather than to inhibit experimentation.

GROWTH AND STORAGE OF BACTERIAL HOST STRAINS
The commonly used hosts for the growth of phage λ are derivatives of Escherichia coli K12. Those few strains to which particular reference is made in this paper are given in Table 1. All known genes of E. coli K12 and their symbols are listed by Bachmann and Low (1980).

For the titration and propagation of λ, bacteria are best grown in a rich medium, such as tryptone broth, with aeration at 37°C, and used when they have reached a...