A reexamination of the Aberrant Ratio phenomenon

The report of aberrant ratios for various marker genes in the advanced progenies of maize plants infected as seedlings with barley stripe mosaic virus (G. F. Sprague and H. H. McKinney, 1971, Genetics 67:533) has elicited considerable interest among geneticists. Attempts have been initiated in several laboratories to investigate the phenomenon further. In a recent paper, R. G. Samson et al. (1979, Genetics 92:1231) reported that the results of their investigations of a Sprague Aberrant Ratio (AR) A/a* stock [a stock supposedly homozygous dominant for all the aleurone factors other than A and in which sib matings in either direction between plants grown from colored seeds (presumed to be A/a) and plants from colorless seeds (presumed to be a/a) yielded an excess of colorless kernels over the 50% expected in such matings] suggested that such stocks might be heterozygous at another locus necessary for aleurone color. They presented no data verifying this hypothesis, however. Our data show by several types of evidence including direct tests of such stocks that two A/a* stocks are indeed heterozygous at other color factor loci. It should be noted that after the initial identification of an aberrant ratio in a self-pollinated plant (e.g., an excess of colorless kernels in a plant of the presumed genotype A/a, A2/A2, C/C, C2/C2, R/R) Sprague and McKinney maintained the AR stocks by sib-matings between plants from colored kernels and plants from colorless kernels. For two other instances of AR, an A*/a stock [in which sib matings between plants grown from colored seeds (presumed to be A/a) and plants grown from colorless seeds (presumed to be a/a) yielded an excess of colored kernels over the 50 percent expected] and an Su/su* stock Lin which sib matings between plants from Su kernels (Su/su) and those from su kernels (su/su) yielded an excess of sugary kernels], other explanations suggest themselves.

In 1977, we received from G. F. Sprague seeds of the cross, A-p/a x a/a [Sprague 1976 441(1) x 442(1)]. This stock gave aberrant ratios of colored (pale) to colorless (an excess of colorless) in sib-matings between plants from colored and plants from colorless seeds. Two types of crosses were made--sib-matings in both directions between plants from colored kernels and plants from colorless kernels and crosses in both directions between our W22 a tester and the plants from colored kernels. The purpose of these crosses was to verify the observations of Sprague and McKinney that (1) there was an excess of colorless kernels in crosses between plants from colored and colorless kernels from the cross of presumed A/a or (a-p/a) x presumed a/a regardless of the direction in which the cross is made; (2) one observes expected proportions (0.5) of colorless kernels when plants from colored kernels (from A/a x a/a) are crossed either as male or female by an a tester; (3) one can recover aberrant ratio stocks after such an outcross.

Table 1. The percentage of colorless kernels observed in various crosses involving Aberrant Ratio stocks
 
Year, row Presumed Genotype Source of Stock Pollinations Made Percent Colorless Kernels
1977, 17687-89 a-p/a Sprague 1976 441(1) x 442(1) Colored (pale) X17690-92 51, 62, 58, 59, 70, 59
      X17695-91 54, 51, 44
1977, 17690-92 a/a Sprague 1976 441(1) x 442(1) Colorless X17687-89 66, 65, 61, 69, 62, 59
1977, 17695-96 a/a tester Nelson 1974 10429 selfed X17687-89 44, 50, 47, 46, 49, 50, 52
Fla 1978, 7824-25 a-p/a 17687 x 695 Colored kernels selfed 22, 46, 27, 43, 28, 21, 22,
        24, 24, 47, 26, 46, 269 25,
        43, 26, 29, 269 43, 29, 42
Fla 1978, 7829 a-p/a 17695 x 687 Colored kernels selfed 26, 27. 25, 43, 24, 40, 33,
        26, 30, 28, 24, 22
1978, 19472-3 a-p/a F7825-2 selfed (47% Colorless) Colored kernels selfed 21, 21, 45, 40, 29, 27, 29,
        0, 40. 46. 46, 41, 43, 44,
        42, 27, 0
1978, 19475-6 a-p/a F7829-6 selfed (40% Colorless) Colored kernels selfed 18, 42, 27, 28, 25, 22, 24,
        30, 43, 23, 0, 0, 27, 45,
        24, 20, 21, 43, 28, 40, 46,
        25, 48, 23, 0. 0

Table 1 records the results of investigations made with this Aberrant Ratio stock from Sprague. The sib crosses between plants grown from colored kernels and those grown from colorless kernels did yield an excess of colorless kernels in most crosses, and crosses of the plants from colored kernels times our a tester did give approximately 50% colorless kernels. However, when colored kernels from this cross (presumed A/a x a tester) were planted in Florida (1978) and the plants selfed, plants giving an excess of colorless were found to occur frequently (Table 1, lines 4 and 5). The percentages of colorless kernels in various plants showed a distinctly bimodal distribution with means of 25.8 and 43.7 for the two classes.

These progenies were followed for one more generation by planting colored kernels from two self-pollinated plants that gave high percentages of colorless kernels in Florida and self-pollinating the resulting plants. The results are given in Table 1, lines 6 and 7. There was a trimodal distribution with respect to the percentages of colorless kernels--those ears with no colorless kernels; those distributed about 25% colorless kernels (x = 24.7%); and those distributed about a modal value in the low 40's (x = 43.4%). By this time, it was clear that all observations could be explained if the original accession from Sprague were, in addition to being heterozygous (A/a), heterozygous at another of the complementary loci necessary for aleurone anthocyanin production. The mean values of 43.7% colorless kernels for selfed plants in Florida (1978) and 43.4% colorless kernels in Wisconsin (1978) on the high colorless plants are close to the 43.75% colorless expected when selfing a plant of the constitution A/a; X/x where either a/a/a or x/x/x conditions a colorless seed phenotype. The mean value of 62.7% colorless in the original sib crosses (Table 1, lines 1 and 3, excluding the value of 51% colorless) is close to the 62.5% colorless expected from a cross of a plant from a colored seed (A/a; X/x) times a plant from a colorless seed (a/a; X/x or A/a; x/x) with either a/a/a or x/x/x conditioning a colorless seed phenotype.

Direct confirmation of the genotype is necessary, however, and this was provided in 1979. Colorless seeds from 19473-5 selfed (43% colorless) were planted, and the resulting plants crossed onto all the color testers. The five plants proved to be uniformly A2/A2; C2/C2; R/R but to be of various constitutions (A/A, A/a, a/a) at the A locus and homozygous recessive c/c at the C locus. A second test demonstrated that colorless seeds within a progeny derived from a cross of the original AR stock (a-p/a) times our a tester could be colorless for reasons other than their genetic constitution at the A locus. Plants from colorless seeds of 19473-6 selfed (44% colorless) were pollinated by our a tester. Of 68 plants thus tested, 38 were a/a, 20 were A/a, and 10 were A/A. Therefore, it is clear that seeds within this progeny could be colorless without being a/a and that alleles at a second color factor locus were segregating. We already know that that locus is C in this line of descent.

A second AR stock was received from Sprague in 1979. This was A/a* x a/a [Sprague 1978 211(1) x 210(1)]. The colored and colorless seeds were planted in adjacent rows, and the plants from the colored seeds were either selfed or crossed reciprocally in paired pollinations with the plants derived from colorless seeds. The results are presented in Table 2. At the same time, 4 plants from colored seeds were used as pollinators onto all color testers. These four plants were all shown to be A/a, A2/A2, C/C, C2/c2, and R/R.

Table 2. The results from sib-matings between plants from colored and colorless seeds derived from A/a* x a/a, Sprague 1978, 211(1) x 210(1), or self-pollinations of plants from colored kernels
 
Year, row Presumed Genotype Source of Stock Pollinations Made Percent Colorless Kernels
1979, 21551 A/a 211(1) x 210(1) Colored kernels - 1 selfed 40
      - 3 x 552-9 74
      - 4 selfed 44
      - 5 x 552-3 76
      - 8 x 552-6 65
      - 9 selfed 42
      -10 x 552-7 57
      -11 x 552-8 75
      -13 x 552-1 71
      -15 x 552-2 59
1979, 21552 a/a 211(1) x 210(1) Colorless kernels - 1 x 551-13 72
      - 2 x 551-15 57
      - 3 x 551-5 75
      - 6 x 551-8 66
      - 7 x 551-10 62
      - 8 x 551-11 78

With the knowledge that alleles at two color factor loci are segregating in the tested plants from colored seeds, it is clear that the colorless seeds from a sib mating might be a/a, C2/c2; a/a, c2/c2; or A/a, c2/c2. This enables one to interpret the results of the sib-matings given in Table 2. These fall into two groups--one with a mean of 61% colorless kernels and one with a mean of 74% colorless kernels. The group with the lower percentage of colorless kernels (x = 61%) almost certainly represents crosses of A/a, C2/c2 plants with plants that are A/a, c2/c2 or a/a, C2/c2. In such crosses, 62.5% of the kernels are expected to be colorless. The second group of crosses (x = 74% colorless kernels) are matings between plants that are A/a, C2/c2 and those that are a/a, c2/c2. In these crosses, 75% of the kernels are expected to be colorless.

A subsection of this progeny was followed in 1980. The colored and colorless kernels from 21551-11 x 21552-8 (75% colorless kernels) were planted in adjacent rows. The plants from the colored kernels were crossed onto the color factor testers, self-pollinated, and used as males onto plants from the colorless kernels in a few instances. The results are given in Table 3. All plants tested were A/a, C2/c2. When selfed, the percentages of colorless kernels were close to the expected 43.75%. When crossed to their colorless sibs, the percentages of colorless kernels were close either to the 62.5% colorless expected if the colorless seed were A/a, c2/c2 (or a/a, C2/c2) or to the 75% colorless kernels expected if the colorless seed were a/a c2/c2.

Table 3. The genotypes and pollination results of plants grown from colored kernels from 21551-11 x 21552-8 (75% colorless kernels)
 
    Pollination Results (% Colorless)
Year, row Genotype(a) Selfed on 23266 (b)
1980, 23265 - 1 A/a ----- C/C, C2/c2, R/R 43 --
- 5
A/a, A2/A2, ---, C2/c2, R/R 46 61
- 8
A/&, A2/A2, C/C, C2/c2, R/R -- 62
-11
A/a, A2/A2, C/C, C2/c2, R/R 44 77
-12
A/a ----- C/C. C2/c2. R/R -- 61
-13
A/a ----- C/C, C2/c2, R/R 45 75

(a) A blank means that the genotype at that locus is unknown.

(b) 23266 contained sib plants grown from the colorless seeds of 21551-11 x 21552-8.

In summary, two Aberrant Ratio stocks giving higher than expected percentages of colorless kernels in sib-matings between plants of the presumed genotypes A/a and a/a were followed for several generations. In each case, the stock was found to be heterozygous at a second color factor locus (C/c in one instance and C2/c2 in the second). The excess of colorless kernels results from segregation of alleles at a second color factor locus. It is unnecessary to postulate any anomalous genetic behavior in these stocks.

Two other Aberrant Ratio stocks have also been received from Sprague and investigated to some extent. The existence of anomalous ratios has been confirmed and the investigations have been sufficient to show that the basis(es) is(are) different than that in the two cases already discussed. The first stock is one in which sib-matings of plants from colored kernels (presumed to be A/a) with plants from colorless kernels (presumed a/a) produce an excess of colored kernels over the 50% expected. One such stock was received from Sprague [1976 433(5) x 434(9)]. It has been most difficult to obtain satisfactory seed sets on this stock so that no adequate data have been collected. In 1979, a plant from a colorless seed crossed as a female with a plant from a colored seed produced 50% colored seeds. Of two plants from colored seeds crossed as females with plants from colorless seeds, one gave 61% colored kernels and the other 51% colored seeds. Although an excess of colored kernels can result from segregation at two color factor loci given the requisite genotypes, that does not account for the results here. In 1980, a number of plants from the colored kernels of the cross that in 1981 produced 61% colored kernels were tested for their allelic constitution at the color factor loci. All such plants were A/a, A2/A2, C/C, C2/C2, and R/R. It is possible that a gametophyte factor linked to A accounts for the excess of colored kernels in sib-matings in this stock. There is a gametophyte factor, Ga7, located 17 map units distal to the a locus. The possibility of the involvement of this or another gametophyte factor previously unidentified should be ruled out before proceeding to alternative suggestions.

Another Aberrant Ratio stock is one in which sib-matings between plants from Su/su kernels and su/su kernels give an excess of sugary kernels. In their 1971 paper (Genetics 67:533-542) Sprague and McKinney suggest that they had found stocks giving aberrant ratios for sugary without supplying any data. In 1979, such an AR stock was received from Sprague [1978, 208(2) x 209(1)]. This was again a sib-mating (Su/su x su/su). The nonsugary (Su/su) kernels were planted and the plants selfed as well as crossed as males to Golden Cross Bantam. The results are presented in Table 4. It should be noticed that contrary to the observations with the (A/a*) AR stocks giving an excess of colorless kernels that outcrosses to a control stock (Sprague and McKinney) or our a tester (Table 1) showed expected ratios (i.e., 50% colorless), outcrosses to a sugary hybrid not previously involved in these crosses still produced an excess of sugary kernels. However, the excess over expectations is greater in the self-pollinations than in outcrosses.

Table 4. The percentages of sugary kernels when plants from the nonsugary kernels from Sprague 1978, 209(2) x 209(1) were self-pollinated or crossed onto Golden Cross Bantam
 
 
Percent Sugary Kernels
Year, row Selfed Onto GCB
1979, 21522 - 1
38 55
- 2
40 --
- 3
45 --
- 4
34 58
- 5
37 56
- 7
27 59
-11
36 64
-12
39 --
-13
38 67
-14
39 61

In 1980, nonsugary kernels from 21522-5 selfed (an Su/su plant that gave 37% sugary kernels when selfed in 1979) were planted. The resulting plants were pollinated by pollen from Golden Cross Bantam (GCB) Hybrid plants and crossed as males onto GCB plants. The results are recorded in Table 5. The data are somewhat incomplete in that both crosses are not available for all plants. It is clear, however, that when the Su/su plants from the AR stock are crossed as females by the su/su GCB hybrid, the percentages of sugary kernels observed in the various ears group about the expected 50%. The reciprocal crosses where the Su/su plants were used as males show marked deviations from 50% suggesting that a gametophyte factor could be involved. There is a gametophyte factor locus, Ga1, on the 4th chromosome linked to the su locus which could produce excess sugary kernels when plants of the constitution Ga su/ga Su were self-pollinated or used as males on a Ga su/Ga su tester. No excess sugary kernels would be expected if Ga su/ga su plants were pollinated by an su/su tester. However, this locus does not seem to be implicated for several reasons: (1) Ga pollen grains have a competitive advantage over ga pollen grains only on the silks of plants which are Ga/Ga or Ga/ga. The GCB hybrid is apparently ga/ga, and Ga pollen grains should not have a competitive advantage over ga pollen grains on GCB silks yet marked departures from 50% sugary are observed when GCB plants are pollinated by Su/su plants from the AR stock; (2) if the AR Su/su plants were ga Su/Ga su, they should be capable of pollinating Ga-s/Ga-s plants, and this was not the case in our tests. Using the term "gametophyte factor" in its broadest sense, there are factors on chromosome 4 capable of distorting segregation ratios for sugary without requiring any specific genetic constitution on the female side. W. R. Singleton and P. C. Mangelsdorf (1940, Genetics 25:366-390) reported small pollen (sp) to be 6 crossover units from su. The sp pollen grains rarely function in competition with Sp pollen grains. Carangal (1958, MS Thesis, Purdue University) reported that lethal pollen (lp) is about 14 crossover units from su. No lp pollen functions in competition with Lp pollen, and the lp pollen cannot be distinguished from Lp by size nor any stain tested. Thus, there is considerable precedent for gametophyte factors on chromosome 4 capable of distorting sugary percentages when pollen from a Su/su plant is used to pollinate an su/su tester if the Su/su plant were Su sp/su Sp or Su lp/su Lp.

Table 5. The percentages of sugary kernels produced when plants from nonsugary kernels (Su/su) of 1979 21522-5 x are crossed as males and females with GCB plants
 
  Percent Sugary Kernels  
Year, row XGCB on GCB
1980, 23397 - 3 -- 65
- 5 -- 67
- 8 42 --
- 9 51 --
-10 47 58
-12 53 62, 63
-13 52 70
-14 53 --

As in the previous cases of AR discussed, the deviation from expectation with the Su/su* stock may well have a prosaic explanation.

I appreciate the cooperation of George Sprague in supplying the various AR stocks used.

Oliver Nelson


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