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Cloning and Expression of the pnd Gene of R16: Determination of Transcriptional Direction and Evolutionary Analysis
Author(s) -
Sakikawa Tomi,
Akimoto Shigeru,
Ohnishi Yoshinari
Publication year - 1985
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.1985.tb00882.x
Subject(s) - biology , cloning (programming) , computational biology , expression (computer science) , genetics , evolutionary biology , microbiology and biotechnology , computer science , programming language
The gene p romoting nucleic‐acid degradation (pnd) of IncB plasmid R16 was cloned into the vector plasmid pACYC177. The pnd gene was found to be located on a 0.55‐kilobase (kb) Alu I‐ Pst I fragment by constructing subclones carrying various portions of the initially cloned fragment. The direction of transcription of the pnd gene was determined by inserting the gene in both orientations into the lacZ′ gene of the plasmid pUR222. In the recombinant plasmid pCM2, transcription of the pnd gene was controlled by the lac promoter region. Addition of cAMP at 42 C resulted in rapid degradation of stable RNA in cells harboring pCM2. In contrast, no RNA degradation was observed in cells harboring pCM14, which has the same insert as pCM2 but in the opposite orientation. The equivalent gene, pnd of IncIα plasmid R483, has previously been cloned, and a detailed restriction map of the region has been constructed (Akimoto, S., and Ohnishi, Y. 1982. Microbiol. Immunol. 26 : 779–793). We constructed a detailed restriction map of the pnd region of R16 and compared it with that of R483. Restriction analyses revealed a similar structure in these two pnd regions. The results suggest that the pnd genes of R16 and R483 have a common evolutional origin.