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Nucleic Acids Research, 1987, Vol. 15, No. 15 5985-6005
© 1987

Articles

Restriction endonucleases for pulsed field mapping of bacterial genomes

Michael McClelland, Robert Jones*, Yogesh Patel and Michael Nelson

Department of Biochemistry and Molecular Biology, University of Chicago 920 E. 58th St., Chicago, IL 60637, USA

Received May 5, 1987. Revised July 6, 1987. Accepted July 6, 1987.

Fundamental to many bacterial genome mapping strategies currently under development is the need to cleave the genome into a few large DNA fragments that can be resolved by pulsed field gel etectrophoresis. Identification of endonucleases that infrequently cut a genome is of key importance in this process. We show that the tetranucleotide CTAG is extremely rare in most bacterial genomes with G+C contents above 45%. As a consequence, most of the sixteen bacterial genomes we have tested are cleaved less than once every 100, 000 base pairs by one or more endonucleases that have CTAG in their recognition sequences: Xba I (TCTAGA), Spe I (ACTAGT), Avr II (CCTAGG) and Nhe I (GCTAGC). Similarly, CCG and CGG are the rarest trinucleotides in many genomes with G+C content of less than 45%. Thus, Sma I (CCCGGG), Rsr II (CGGWCCG), Nae I (GCCGGC) and Sac II (CCGCGG) are often suitable endonucleases for producing fragments that average over 100,000 base pairs from such genomes. Pulsed field gel electrophoresis of the fragments that result from cleavage with endonucleases that cleave only a few times per genome should assist in the physical mapping of many prokaryotic genomes.

*Present address: Department of Biological Sciences, University of Southern California, University Park, Los Angeles, CA 90089, USA


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