DISTRIBUTION AND CHARACTERIZATION OF MUTATIONS INDUCED BY NITROUS-ACID OR HYDROXYLAMINE IN THE INTRON-CONTAINING THYMIDYLATE SYNTHASE GENE OF BACTERIOPHAGE-T4

the detailed distribution and characterization of 51 hydroxylamine (HA )-induced and 59 nitrous acid (NA)-induced mutations in the intron-con taining bacteriophage T4 thymidylate synthase (td) gene is reported he re. Mutations were mapped in 10 regions of the td gene by recombinatio nal marker rescue using plasmid or M13 subclones of the td gene. Phage crosses using deletion mutants with known breakpoints in the 3' end o f the td intron subdivided HA and NA mutations which mapped in this re gion. At least 31 of the mutations map within the 1-kb group I self-sp licing intron. Intron mutations mapped only in the 5' and 3' ends of t he intron sequence, in accordance with the hypothesis that the 5' and 3' domains of the T4 td intron are essential for correct RNA splicing. RNA sequence analysis of a number of mapped td mutations has identifi ed two intron nucleotides and one exon nucleotide where both HA- and N A-induced mutations commonly occur. These three loci are characterized by a GC dinucleotide, with the mutations occurring at the cytosine re sidue. Thus, these data indicate at leas three potential sites of both HA- and NA-induced mutagenic hotspot activity within the td gene.