Chemically induced paramutation at the R locus in maize.

The R gene in maize, conditioning anthrocyanin formation in seed and plant, invariably is changed from a darkly pigmenting to a weakly pigmenting (repressed) form in the progeny of heterozygotes carrying the stippled (Rst) allele. The repressed forms of R are gametically transmitted, and are referred to as paramutant (R') alleles.' 3 The evidence presented in this report shows that R' alleles are highly sensitive to heritable changes in level of pigmenting action in response to treatment of seed with diethyl sulfate or ethyl methanesulfonate. Mutant sectors were detected in more than 90 per cent of the treated Ml plants in some cases, showing that the frequency of mutagen-induced change may approach that of R in RR't plants. The frequency of change in level of R' action induced by these chemicals is far higher than that characteristic of mutation in general, and suggests that chromosomal components other than genes in the conventional sense are involved. Materials and Methods.-Gene symbols: The standard Rr allele conditions colored aleurone and red plant. It gives self-colored aleurone in two or three doses (RrRrRr or RrRrr) and darkly mottled aleurone in single dose (Rrrr). R69 is a mutant from standard Rr, equivalent to the latter in aleurone pigmenting action but conditioning green (no anthocyanin), rather than red, plant. The rr and rg genes give colorless aleurone and red and green plants, respectively. Stippled (Rat) is a strongly paramutagenic factor which also causes aleurone spotting. The strongly repressed allele, Re,', used initially in testing sensitivity of'paramutant R' forms to chemical mutagens, was isolated among the descendants of R6' R69' plants derived originally from a self-pollinated R6s6Rt individual. All these alleles had previously been introduced into a common, highly inbred strain known as W22. Chemical mutagen treatments: Two well-known chemical mutagens, which act as monofunctional alkylating agents, diethyl sulfate (DES) and ethyl methanesulfonate (EMS), were used in these experiments. Because DES is subject to rapid hydrolysis in aqueous solutions, the preparations of it were changed every 100 minutes during seed treatments. Unless otherwise noted, each treatment involved soaking 150 dormant seeds in 500 ml of the aqueous mutagen solution at room temperature (ca. 230C). The treated seeds were twice rinsed and soaked in distilled water for 6 to 12 hours, then planted immediately. Scoring aleurone pigmentation: Standard procedure in evaluating the pigmentproducing action of an R allele is to apply pollen from the plant to be tested to the silks of an rr individual. The resulting seeds, carrying R in single dose, are scored by matching them with a standard set of six kernels, defining seven color classes, ranging from colorless (class 1) to fully pigmented (class 7). Usually the mean score for 40 kernels from a testcross ear was used as the measure of aleurone pigmenting action of the R allele under test. In some. experiments entire testcross ears were matched against a graded series of standard ears defining five aleurone color levels ranging from colorless (class 1) to darkly mottled (class 5).