Cloning the gene for ribonuclease E, an RNA processing enzyme

Abstract

A transducing bacteriophage lambda Ch25rne+, which codes for ribonuclease E of E. coli, has been isolated. To achieve this a random library of Escherichia coli HindIII fragments was cloned in the lambda Charon 25 vector (prepared in F.R. Blattner's laboratory), and lambda Ch25rne+ was selected by its ability upon lysogenization to enable a temperature-sensitive (ts) rne-3071 mutant to grow and to exhibit normal RNA processing at the nonpermissive temperature of 45 degrees C. The level of RNase E was doubled in an rne+ strain lysogenized with lambda Ch25rne+. lambda Ch25rne+ directs the synthesis of a polypeptide of 71 000 m.wt., which is the size of RNase E. Restriction analysis and electron micrography of heteroduplexes suggested that the size of the host DNA insert is about 1.9 kb.