Enrichment of genes and location of mutations in cloned DNA fragments of Streptococcus pneumoniae

Abstract

Genetic transformation of the naturally transformable bacteria Streptococcus pneumoniae with chromosomal DNA cleavaged with various restriction enzymes allowed the determination of the residual biological activity of the restricted DNA. Electrophoretic fractionation of the digested DNA followed by transformation with the eluted DNA bands led to the identification of the sizes of the fragments which contained activity for 3 chromosomal markers, viz. nov-1, str-41 and, as a control, malM-594, for the restriction enzymes EcoRI, BglI and HindIII. In addition, a 3.1-kilobase pairs (kb) pneumococcal insert containing the nov-1 mutation has been cloned in pBR328. Genetic and restriction analysis of the recombinant plasmid allowed the mapping of this mutation within a Sau3A-BglI chromosomal fragment of 0.5 kb. © 1987.