Sadao Ichikawa

Somatic mutations in Tradescantia stamen hairs exposed to ethyl methanesulfonate

Abstract Young inflorescences of Tradescantia (BNL clone 02) heterozygous for flower color were treated with 0 (water), 0.1 or 0.5% EMS solutions for 2, 4, 8 or 16 hr. The frequency of somatic pink mutations in the stamen hairs scored 10–18 days after treatments was higher with the higher concentration of EMS and increased with lengthening duration of the treatment. The mutation frequency was also found to be clearly a function of the EMS dose in %·hr. The frequency of mutation increases with EMS dose; on a log-log graph the slopes were 1.37 or 1.31, when the mutation frequency was expressed as the number of pink mutant events per 10 3 hairs or per 10 5 cell divisions, respectively. That the slopes are steeper than 1 suggests the involvement of two-break deletions, but the possibility of insufficient penetration of EMS in cases of shorter treatments cannot be ruled out. The mutation frequencies observed were somewhat comparable to those induced by 0.25 to 10 rad of sparsely ionizing radiations at various dose rates. (1 − 4, 6, 16, 17)

Somatic mutation frequencies in the stamen hairs of stable and mutable clones of tradescantia after acute gamma-ray treatments with small doses ☆

Abstract Young inflorescences of two different Tradescantia clones heterozygous for flower and stamen-hair color, one stable (KU 9) and the other spontaneously mutable (KU 20), were irradiated acutely with small doses (approx. 3 to 50 R) of 60 Co gamma-rays. Somatic mutation frequencies from blue to pink in the stamen hairs scored on post-irradiation days 10 to 16 increased essentially linearly with increasing gamma-ray dose in both clones. Despite about a 5-fold difference in spontaneous mutation frequency per hair found between the two clones, the dose-response curves of pink mutations determined were similar to each other, giving average mutation frequencies of 1.51 and 1.41 pink-mutant events per 1000 hairs per R for KU 9 and KU 20, respectively. These frequencies are comparable to earlier results obtained from acute irradiation treatments of other clones with higher doses. The doubling dose of pink mutation (the radiation dose making the mutation frequency double the spontaneous level) was calculated to be 2.09 R for KU 9, and this low doubling dose must be given full attention. On the other hand, the doubling dose for KU 20 (calculated to be 10.4 R) is of questionable value, being greatly subject to change because of the diversely variable spontaneous mutation frequency of this clone.

RADIOSENSITIVITY OF A TRIPLOID CLONE OF TRADESCANTIA DETERMINED IN ITS STAMEN HAIRS.

Somatic mutations from blue to pink were induced by 60 Co gamma rays in the stamen hairs of KU 9 clone (3x) at a rate of 1.64 × 10 −3 pink mutant events per hair per R or 2.32 × 10 −4 pink mutant cells per hair cell per R. A D 0 value of 171 R was determined for the terminal meristematic cells of the stamen hairs. This D 0 value was compared with ten other D 0 values obtained earlier from seven different 2x, 4x or 12x clones, and it was reconfirmed that higher radioresistance of higher polyploids is best explained by their smaller chromosome volumes rather than by genetic redundancy. The effects of dose rate on somatic mutation rate and loss of reproductive integrity were also clearly demonstrated. Pink mutant cells were found to have a lower reproductive ability on the average than non-mutant cells.

Variation of spontaneous mutation frequency in Tradescantia stamen hairs under natural and controlled environmental conditions

Abstract Considering the lack of information about the variation in spontaneous mutation frequency, especially due to different genotypes and under different environmental conditions, a long-term scoring of spontaneous pink somatic mutations was carried out in the stamen hairs of three different clones of Tradescantia heterozygous for flower and stamen-hair color, keeping them under controlled or natural conditions. One of the clones, KU 20, was found to be highly mutable and also unstable showing about a 23-fold difference in the spontaneous mutation frequency depending upon the environmental condition. The other two clones, KU 7 and KU 9, were rather stable and much less mutable than KU 20. High negative correlations were found between spontaneous mutation frequency and temperature in clone KU 20, i.e., the lower the temperature (in the range of about 18 to 28°C) the higher was spontaneous mutation frequency, and this clone was found to be especially sensitive to temperature variation. A significant (at the 5% level) negative correlation between spontaneous mutation frequency and temperature was also found in KU 7, and a similar, but not significant tendency was observed in KU 9. The spontaneous somatic mutation frequency in older inflorescences of clone KU 20 was significantly lower than that in the younger inflorescences.

Radiobiological studies in plants—XI: Further studies on somatic mutations induced by X-rays at the al locus of diploid oats*

Abstract The effects of X-ray dose rate, fractionated exposure, and DNP treatment on somatic mutation frequency were studied using a mutant strain of diploid oats heterozygous for the albino gene. Various X-ray treatments were conducted on both the dry and 24-hr pre-soaked seeds. Higher dose rates were more effective in inducing mutations than lower dose rates. Fractionated exposure induced less mutations than non-fractionated exposure. Similar fractionated exposure but accompanied with a DNP treatment had about the same efficiency as the non-fractionated exposure. Consequently, it was concluded that many of the induced somatic mutations were induced by chromosomal deficiencies. The number of white stripes per leaf decreased with the progress of leaf order. This phenoemenon appears to be due to the difference in numbers of the primordial or initial cells at the time of irradiation. A similar relationship was observed between the average ratio of white stripe width to leaf width and leaf order. It seems possible to consider that a considerable elimination of mutant cells occurred in the course of leaf development. Distinct white stripes were observed in leaf 3 and upper leaves but never in leaf 1 and 2 after irradiations of dry seeds. However, such distinct stripes were also observed in leaf 2 after irradiations of 24-hr pre-soaked seeds. This phenomenon is discussed giving two possible explanations.

RADIOBIOLOGICAL STUDIES IN PLANTS. X. MUTATION RATE INDUCED BY IONIZING RADIATIONS AT THE AL LOCUS OF SAND OATS

Abstract In order to elucidate a relationship between radiation dose rate and somatic mutation rate, a mutant line of sand oats known as green inconstant, heterozygous for plant color gene (+/ al ), was used in the present study. Dry seeds of the green inconstant were exposed to neutrons from a reactor and gamma rays from Co 60 or Cs 137 . All of the doses were simultaneously measured with several appropriate dosimeters at every irradiation treatment (Tables 1 and 2). Distinct white stripes were found in leaf 3–6 of seedlings, but those in leaf 1 and 2 were too small and indistinct to score. All induced stripes in leaf 3–6 extended from the base to the apical part of leaf blades and remained the same width throughout. The number of white stripes per leaf and the average ratio ( p %) of white stripe width to leaf width were used as criteria of the somatic mutation frequency by radiation treatments. It is evident that the frequencies of white stripes increase with dose in each leaf, and the dose-response relationship does not appear to be linear, especially in the gamma-ray treatments (Figs. 7 and 9). Dose-rate effects were also observed, the higher dose rate being more effective. These responses are explicable by assuming a combination of one-hit and two-hit events. On the other hand, the frequencies of white stripes was reduced successively from leaf 3 to 6 (Figs. 6 and 8). This phenomenon seems mainly to be due to the developmental pattern of plant, and partly to elimination of mutant cells. Based on a consideration of leaf ontogeny it is finally concluded that the average ratio ( p %) of white stripe width to leaf width in leaf 3 shows more approximate value of the mutation rate at the al locus than those in leaf 4 to 6 (Table 4).

A developmental study of diploid oats by means of radiation-induced somatic mutations*

Abstract The developmental behaviour of radiation-induced mutant cells in X 1 plants was studied using a strain of diploid oats heterozygous for the albino gene, al . Dry and 24-hr pre-soaked seeds were irradiated with X-rays, γ-rays and thermal neutrons. Distinct white stripes were observed in leaf 3 and upper leaves when dry seeds were irradiated, but from leaf 2 when pre-soaked seeds were treated. The different response in the pre-soaked seeds might be caused by the radiation-induced loss of reproductive integrity of most of the sensitized leaf 2 primordial cells followed by reorganization of the primordia in the embryos. The number of white stripes per leaf decreased with the progression of leaf order. From the relative widths of the stripes, the number of primordial or initial cells for each leaf in the mature embryos was estimated to be about 30 or less for leaf 3 and at least three to five for leaf 4 to 6. Since the stripes in leaf 3 from the treated dry seeds were induced independently from those in the upper leaves, leaf 3 seems to have been derived from an already differentiated primordial cell group, although such a group could not be detected histologically. The white stripes were more frequently observed near the centre of the leaf blade rather than in the marginal part with wider stripes tending to appear in the marginal region, indicating a differential development in the lateral direction. The tillers on the same orthostichy tended to behave similarly with regard to the appearance of the white stripes or sectors. The present study demonstrates that mutations having an appropriate phenotype induced in somatic tissues provide a useful means for detailed study of organogenesis in higher plants.

TWO KINDS OF CHROMOSOME REJOININGS IN X-RAYED TWO-ROWED BARLEY

Abstract In order to determine whether or not some rejoining of broken ends of chromo-somes occurs within a matter of seconds after irradiation, X-ray irradiation was performed on the seeds of two-rowed barley presoaked for 24 hr, changing the exposure and the dose rate or fractionating the exposure. All the exposures or dose rates used were measured simultaneously with or immediately before the treatments, and all the exposure times and the intervals in fractionation experiments were accurately measured. In each lot, the frequency of induced dicentric bridges was scored at anaphase of the first mitotic division in 2000 to 6700 cells fixed 48 hr after the start of presoaking. All the experiments were carried out at 20 ± 1 °C. The frequency of dicentric bridges increased with exposure, giving a curve close to square-of-exposure (Figs. 2 and 4), and effects of dose rate and of fractionation were clearly observed (Fig. 1 and Table 3). It was also made clear that there are two kinds of X-ray induced chromosome breaks, one of which can rejoin within an extremely short period such as 20 to 30 sec after irradiation, and another can rejoin after a longer time such as 15 to 30 min (Fig. 3). The latter breaks were induced about 20 per cent more frequently than the former breaks. From the experiment using DNP, it was presumed that the energy required for rejoining is supplied from oxidative phosphorylation.

VARIATION IN SOMATIC CHROMOSOME NUMBER FOUND IN RADIATION-INDUCED MUTANTS OF CHRYSANTHEMUM MORIFOLIUM HEMSL. CV. YELLOW DELAWARE AND DELAWARE.

Abstract Young plants of Chrysanthemum morifolium CV. Yellow Delaware and Delaware were irradiated chronically with 60 Co gamma rays in the gamma field. The total exposures were 5, 10 and 20 kR. Many whole-type sports were produced after irradiation, thus the method of internal disbudding by radiation treatments demonstrated earlier, (18) was confirmed to be efficient in enlarging mutation sectors. Cytological examination of 19 mutant and 12 apparently normal shoots produced after irradiation revealed that the somatic chromosome numbers ranged from 49 to 55 with an exceptional doubled number of 110. Some of the shoots were chimeric cytologically. None of the shoots examined had higher chromosome numbers than the control plants (2 n = 55) except for the 110-chromosome shoot. More reduction of chromosome number was caused with higher exposures. Reduction of chromosome number was also correlated with the type of mutation and with the size of inflorescence.