FREQUENCY OF RECONSTITUTION OF THE VARIEGATED PERICARP ALLELE IN MAIZE

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FREQUENCY OF RECONSTITUTION OF THE VARIEGATED PERICARP ALLELE IN MAIZE1

ELWIN R. ORTON?

Department of Genztics, University of Wisconsin, Madison

Received July 6, 1965

SELF-RED pericarp

(P)

in maize arises as a mutant from variegated pericarp

( P r r M p ) by the transposition of Modulator ( M p ) away from the P locus, where it represses Prr action, to another chromosome site. Evidence summarized in the preceding article (ORTON and BRINK 1966) showed that occasionally the variegated allele is reconstituted in a Prr plant hemizygous f o r such a transposed Modulator ( t r - M p ) by transposition of M p back to the P locus. The data now to be presented demonstrate that the frequency with which the variegated allele is resynthesized in such cases is related to the proximity of tr-Mp to the P locus. This finding is in accord with expectations based on the observations of VAN SCHAIK and BRINK (1959) and GREENBLATT and BRINK (1962) that when Modulator transposes it often moves a short distance within the same chromosome.

MATERIALS A N D METHODS

Twenty-three P r r families representing nine mutant P r r alleles that had arisen independently in sublines of an inbred W23 strain into which a common variegated pericarp allele originally had been introduced were used in this study. The sublines were maintained by recurrent mating of PrrP*or plants to a W23 PtorPwr stock culture (colorless pericarp, red cob). Numerous W23

P r P w r and Pl"rPt"r segregates repr-senting each P r r mutant were grown in a detasseling plot in which a C-Ds tester stock was used as the staminate parent. The tester stock carried Ds in standard position (MCCLINTOCK 1951) o n chromosome 9 and also the dominant aleurone-color factor, C, in a distal location on the same arm. Modulator, regardless of its position in the genome, induces breakage at the site of Ds, resulting in the loss of C in the acentric fragment. Since the pistillate parents were cc, the loss of C gave rise to colorless areas in an otherwise colored aleurone, and so served to test for presence or absence of Modulator in the genome.

The red ears ( P r r W r ) were scored after harvest for number and size of sectors showing a variegated o r nearly colorless phenotype. Seven size classes were used, in accordance with the procedure employed by WOOD and BRINK (1956). The smallest class comprised the sectors extend- ing over 1/32 to 1/16 of the kernel circumference. Class width doubled progressively over the remainder of ths scale up to the top class which included the sectors involving two o r more kernels. The term "sectoring rate" denotes the total number of sectors, regardless of size, per 1000 kernels. Kern21 number was estimated by multiplying the average obtained from the sum of the number of kernels in the longest and shortest double rows o n an ear by the number of double rows present. Most of the sectors werz too small to involve the embryo. Thus, the mutations were not transmitted to the offspring. The sectoring rates given, therefore, are based entirely on phenotypic changes.

Paper No. 1031 from the Department of Genetics, Agricultural Experiment Station, University of Wisconsin, Madison. Present address: Department of Horticulture and Forestry, Rutgers-The State University, New Brunswick, New Jersey.

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18 E. R. ORTON

Pericarp sectors may arise from a variety of causes in PrrPzor plants, such as mutation of

Prr to colorless, a deficiency involving Prr, nondisjunction of the chromosome 1 carrying Prr, or a nuclear factor different from, but acting as a repressor of Prr, like M p . It is to be expected, therefore, that in the PrrPwr stocks there will be a background rate for pericarp sectoring that is unrelated to M p action. The procedures used to estimate the background rate will be described later.

The conventional test f o r linkage between P and M p located on chromosome 1 does not yield an uncomplicated estimate of the value because the results of crossing over and secondary transposition of M p in these stocks are confounded with each other. The transposition rate of M p in W23 medium variegated pericarp plants is of the order of 10%. Two circumstances, however, make the error thereby introduced into estimates of linkage between P and M p considerably less than this amount: (1) Many M p transpositions are to nearby sites on the original chromosome and (2) when M p transposes to another chromosome, the latter would assort in meiosis with the chromosome 1 carrying Prr i n half the cases. Some transpositions, nevertheless, simulate the effect of crossing over between P and M p . The errors thereby introduced into a linkage estimate will be inversely proportioned to the proximity of M p to the P locus, assuming a constant M p transposition rate.

RESULTS

The data on pericarp sectoring frequencies in relation to recombination between transposed Modulator and Prr are assembled in three groups. The results from seven P r r families derived originally from a single self-red kernel on a medium variegated ear are summarized in Table 1. Six other P r mutants resulting from independent mutations of the original

P""

allele are represented by the ten families referred to in Table 2. Pericarp sectoring is associated with the presence of M p in the genome in both these groups.

The results from six additional PrrPor families, based originally on two independently occurring self-red mutants on a single medium variegated ear are brought together in Table 3. It will be seen that in this group of families a factor, or factors, other than M p exerted a major effect on frequency of pericarp sectoring.

The relationship between the seven families referred to in Table 1 is shown be- low:

Backcross generation of PrrPwr plants (recurrent parent = W23 P w r P r )

7 8 9 10

01115 01116

8-9684-

rg53

0955 ---+ 0970-

10956

-

0969

Five of the families in this group, namely, 0953, 0955, 0956, 0970, and 01 115, showed pericarp sectoring rates of the order of 14 per 1000 kernels, and less than 4.5

%

recombination between tr-Mp and Pr, as shown in Table 1 , Family 0969, on the other hand, displayed a sectoring rate of 0.2 per 1000 kernels, and a recombination value between tr-Mp and P r r of 45.4%. The pericarp sectoring rate

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20 E. R. ORTON

common origin; a low sectoring rate was associated with very loose linkage, or possibly independence, of tr-Mp and P" in another family, and also the absence of M p from the genome in still another instance. Evidently a secondary transposition of tr-Mp occurred in the P r r P ' parent of family 0969 in which M p pre- sumably was initially closely linked to P. Elimination of M p from the genome could have occurred at meiosis in the p"P"' parent of family 01116 following crossing over of tr-Mp from the PT to the Pwr chromosome or transposition of tr-

M p to a new site.

Six of the families referred to in Table 2 lacked M p in the genome. The sectoring rate was low in all these cases, and averaged 0.1 per 1000 kernels. The frequency of sectoring was only slightly higher for the M p f plants in family 0966, namely, 0.2, in which 41.8% recombination between Prr and tr-Mp was observed. The remaining three families, 0951, 0952, and 0975, derived from M p + plants in which tr-Mp and P r were more closely linked and showed much higher sectoring rates. It is noteworthy also that within these families pericarp sectoring rates were

4

to 10 times higher in M p + than in Mp- plants.

Within families 0951, 0952, and 0975, however, there is little relationship between sectoring rate and percent recombination between tr-Mp and

P.',

The

M p + plants in families 0952 and 0975, for example, gave sectoring rates of about 11 per 1000 kernels in each case, but the respective tr-Mp and Prv recombination values were widely different from each other, namely, 4.7 and 17.8%. This dis- crepancy is possibly due to overestimation of the amount of recombination between tr-Mp and P r in family 0975. The criterion for presence of M p is patches of colorless aleurone in C-Ds/c-ds/c-ds testcross endosperm resulting from breaks at Ds in response to M p action in the cells. Few colorless aleurone patches occurred among the testcross kernels in family 0975. This irregular effect of Ds in causing C losses could have resulted in classifying some M p + plants as Mp- and so have given an overestimate of the amount of recombination between P r and

tr-Mp.

The six families the results from which are entered in Table 3 illustrate varia- tion in pericarp sectoring that is not associated with M p . These families derive from two independently occurring mutations to self-red pericarp on a single medium variegated ear, 6F-157-1, as summarized in the following diagram:

Backcross generation of P r r P w r plants (recurrent parent = W23 PW'P')

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0948-+

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It will be noted that within each of the four families segregating for M p ,

namely, 0945, 0948, 01 113, and 01 114, the pericarp sectoring rates for the Mp+

and Mp- plants are of similar magnitude. Furthermore, in family 0972 which was Mp- throughout, the sectoring rate was 4.6 per 1000 kernels, a value which approximates the maximum (5.4 per 1000 kernels) observed for the Mp+

plants among the families segregating for Modulator. The seed ear giving rise to family 01113 exhibited a sectoring rate comparable to the most highly sec- tored sib ears in family 0948, whereas no sectors were evident on the seed ear giving rise to family 01114. The selection of these seed ears on the basis of the number of pericarp sectors resulted in the isolation of two sib families differing markedly in the stability of Prr expression. The sectoring rate and percent recombination of tr-Mp and Prr computed from the combined data for these two families are very similar to the respective values for the parent family. It is ap- parent that segregation of some factor other than M p markedly influenced pericarp sectoring in family 0948. Evidently most of the pericarp sectoring shown by the group of related families listed in Table 3 is due to some factor other than

M p . The presence in certain of these families of tr-Mp at a distance of 10 to 15 crossover units from the P locus had little, if any, effect on the frequency of sectoring.

DISCUSSION

The data presented in Tables 1 and 2 show that the frequency of pericarp sectoring in self-red mutants derived from the variegated allele is related to the recombinational distance between P r r and transposed Modulator. In general, increases in pericarp sectoring rates are proportional to the diminution of the map distance between tr-Mp and P r r . The seven families entered in Table 1, originally derived from a single self-red mutant kernel from variegated, provide especially cogent evidence on this relation. Modulator was absent from the genome in one of these families, namely, 01116; the sectoring rate in this case was only 0.1 per 1000 kernels. Low sectoring values were observed also in the relatively few Mp- plants present in families 0953, 0955, 0956, 0970, and 01115 in which tr-Mp and Prr showed less than 5% recombination. The rates observed in all these cases reflect the effects of the residual heredity on P r losses in this stock. The M p + plants in these respective families, on the other hand, showed sectoring rates of 10.7, 14.7, 13.3, 14.4, and 16.5 per 1000 kernels. By contrast, family 0969 showed 45.4% recombination between tr-Mp and Prr. The sectoring rate in the Mp+ plants in this family was of the same order of magnitude as in the Mp- sibs, namely, 0.2 per 1000 kernels.

The evidence from 10 additional self-red families, assembled in Table 2, is similar. Low sectoring rates were observed in the six families that were Mp-

throughout and also in the M p - plants in the four families segregating tr-Mp.

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24 E. R. O R T O N

tr-Mp was evident in the latter instance, and the sectoring rate was only 0.2 per 1000 kernels in the M p f , as well as in the Mp-, segregates.

Family 0975 in this group gave results that did not conform to the general rule that the frequency of sectoring is positively related to the proximity of transposed Modulator to the P r gene. Difficulties were encountered, however, in classifying the plants in this family for presence or absence of M p due to failure of the C-Ds

test to yield an unequivocal expression of C losses in the aleurone tissue. Doubt remains, therefore, that the 0975 results constitute a valid exception to the relationship observed in the other families in the same group.

Evidence 'was presented in Table 3 which demonstrates that pericarp sectoring may be significantly affected by a factor (or factors) that operates quite independently of Modulator in the genome. These results do not contradict the conclusion that sectoring in Prr mutants from variegated pericarp is a function of the proximity of the transposed Modulator to Pr. They merely show that the phenotypic effect of transposition of M p back to the P locus in a mutant Prr plant may be simulated by other seemingly unrelated factors. Parallel examples are already known in maize, one of which is repression of A , gene action in the quite distinct Dt, Ds-Ac, and S p m mutation systems (RHOADES 1938; MCCLINTOCK

1956).

The author wishes to express his gratitude to DR. R. A. BRINK for his kind help in the prepa- ration of the manuscript.

SUMMARY

Reconstitution of the variegated pericarp allele ( P r M p ) in P r mutants hemizygous for transposed Modulator ( t r - M p ) has been demonstrated to occur as a result of transpositions that return M p to the P locus. It was predicted that the frequency of synthesis of the variegated allele in such plants would be positively related to the proximity in the chromosome of tr-Mp to the P locus. Evidence was obtained in the present experiments supporting this hypothesis. One body of seemingly nonconforming data involved uncertainties of classification for presence or absence of M p in the genome. Evidence was obtained showing that pericarp sectoring phenotypically indistinguishable from that resulting from the return of transposed Modulator of the P locus in P r P r plants may arise from the action of heritable factors other than M p . The data in these cases are not contradictory to the main conclusion that the frequency of reconstitution of the variegated pericarp allele in self-red M p + mutants is a function of proximity to the P locus of the transposed Modulator.

LITERATURE CITED

GREENBLATT, I. M., and R. A. BRINK, 1962 MCCLINTOCK, B., 1951

Twin mutations in medium variegated pericarp in maize. Genetics 47 : 489-501.

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25

Reconstitution of the variegated pericarp allele in maize Effect of the Dt gene on the mutability of the a, allele in maize. Genetics Transpositions of Modulator, a component of the ORTON, E. R., and R. A. BRINK, 1966

RHOADES, M. M.. 1938

by return of Modulator to the P locus. Genetics 53: 7-16.

23: 377-395.

VAN SCHAIK, N. W., and R. A. BRINK, 1959

variegated pericarp allele in maize. Genetics 44: 725-738.

Frequency of somatic mutation to self color in maize plants homozygous and heterozygous for variegated pericarp. Proc. Natl. Acad. Sci. U.S. 42:

514-51 9.