Genetics and Cytology of Chromosome Inversions in Soybean Germplasm.
Cytogenetic studies of interspecific hybrids between the cultivated soybean G. max and the wild annual soybean G. soja revealed the presence of one or more paracentric inversions (Ahmad et al., 1977a, 1977b, 1979, 1983). These interspecific hybrids exhibited marked differences in chromosome behavior and fertility depending on parents and temperature. In general, summer-grown plants had a higher frequency of meiotic irregularities than did winter-grown plants (Ahmad et al., 1977b). Plants grown in controlled environment cabinets confirmed the effect of high temperature causing more irregularities in chromosome behavior (Ahmad et al., 1983). Furthermore, differences among cross-combinations involving two G. max parents with two G. soja parents were genotype specific (Ahmad et al., 1983). Hybrids within the perennial wild soybean species have shown the presence of paracentric inversions (Singh and Hymowitz, 1985; Hymowitz et al., 1991).
In a search for chromosome aberrations among cross-combinations of G. max germplasm, Delannay et al. (1982) and Palmer (1985) reported that seven of 626 combinations gave [F.sub.1] hybrids with 10 to 30% pollen sterility. A similar study with G. soja germplasm showed that 20 suspected inversions from 142 cross-combinations gave 10 to 40% pollen sterility (Delannay et al., 1982; Palmer, 1985). The G. soja accessions from Japan and South Korea had a higher frequency of suspected inversions than did accessions from China, Taiwan, and Russia. One G. soja accession from South Korea, PI 407179, has been confirmed to differ from G. max by a paracentric inversion (Harper, 1992). This PI was backcrossed to the cultivar Hark (Weber, 1967) to create a near-isogenic line homozygous for the paracentric inversion (Palmer, 1981, unpublished data).
Sun et al. (1991) have identified paracentric inversions in two Chinese G. max landraces, Wei Da Yu and Sun Wu Xiao Bai Mei. These landraces have been entered into the USDA soybean germplasm collection as PI 597651 and PI 597652, respectively.
Our objective was to determine if the paracentric inversions identified in G. soja PI 407179 and in G. max PI 597651 and PI 597652 were identical.
MATERIALS AND METHODS
The two Chinese G. max landraces, Wei Da Yu and Sun Wu Xiao Bai Mei, were identified previously as differing from other G. max lines by a chromosome inversion (Sun et al., 1991). The two Chinese homozygous inversion genotypes were used directly in cross-pollinations. PI 407179 (G. soja) was identified as differing from G. max by a chromosome inversion (Delannay et al., 1982; Palmer, 1985). The inversion from PI 407179 was backcrossed into cultivar Hark (BC6) to develop a near-isogenic line homozygous for the inversion. Selection for inversion heterozygosity was practiced between each backcross by identifying plants expressing partial pollen sterility. The three homozygous inversion lines (PI 597651, PI 597652, and Hark homozygous inversion from PI 407179) were each crossed to Hark and were intercrossed with each other to create three homozygous inversion x homozygous inversion combinations.
The [F.sub.2] plants from the three homozygous inversion lines x Hark were grown at the Bruner Farm near Ames, IA, which has Clarion and Nicollet loam soils (fine-loamy, mixed, superactive, mesic Typic Hapludoll and fine-loamy, mixed, superactive, mesic Aquic Hapludoll). These plants were used for pollen analyses. The [F.sub.1] plants used for meiotic analyses were grown in the glasshouse at Ames, at 24 [+ or -] 2 [degrees] C (dark) and 29 [+ or -] 2 [degrees] c (light). Day length was 14 h of light and 10 h of darkness. The photosynthetic irradiance range was 270 to 500 [micro]mol photon [m.sup.-2] [s.sup.-1]. At Gongzhuling, China, the [F.sub.1] plants used for meiotic analyses were grown in 5-L pots in fine-silty, mixed, superactive mesic Typic Hapludoll soil outside during the summer under natural photoperiod. Temperatures were 17 [+ or -] 2 [degrees] C (dark) and 22 [+ or -] [degrees]2C (light).
Fertile and heterozygous chromosome inversion plants were identified by squashing anthers in an aqueous solution of I2KI (Jensen, 1962, p. 203). Anthers from fertile plants gave densely stained reddish-brown pollen grains, whereas anthers from heterozygous inversion plants gave both well-stained pollen grains and aborted (smaller and lightly stained) pollen grains. Five hundred pollen grains were counted from each plant examined.
Immature floral buds for meiotic analyses were collected from fertile and heterozygous inversion glasshouse-grown plants (Ames, IA) during the first 6 h of the light cycle. The buds were fixed immediately in either 3:1 absolute ethanol/ chloroform saturated with ferrous acetate or 6:3:2 absolute ethanol/chloroform/propionic acid saturated with ferrous acetate. The buds were fixed for a minimum of 24 h, washed twice with 70% ethanol, and stored at 4 [degrees] C. For microscopic examination, the buds were rinsed with distilled water, placed in 45% propionic acid, and stained with propionocarmine. The microsporocytes were scored for frequencies of bridges and fragments. Pollen grains and anther squashes were viewed on a Zeiss Standard WL Research microscope (Carl Zeiss, Thornwood, NY) using an MC63 Photomicrographic Camera Attachment (Brinkmann Instruments, Des Plaines, IL) and Kodak Techpan film (Eastman Kodak Co., Rochester, NY).(1)
At Gongzhuling, China, all five cross-combinations were evaluated cytologically. Immature floral buds for meiotic analyses were collected and fixed similarly as in Ames, IA. Pollen grains and anther squashes were viewed on a Leitz orthoplan microscope (Leica, Deerfield, IL) with a 35-mm camera attachment and Kodak Techpan film.
RESULTS AND DISCUSSION
The intercrosses of the two Chinese landraces gave completely fertile pollen for all the [F.sub.1] and [F.sub.2] plants (Table 1). These two landraces have the identical chromosome inversion, consistent with the observations of Sun et al. (1991).
The three homozygous inversion lines were crossed to cultivar Hark (N N) and pollen was classified from [F.sub.1] and [F.sub.2] plants (Fig. 1A and 1B). The cross of Hark (N N) with Hark homozygous inversion (In In) gave [F.sub.1] plants with about 25% pollen sterility (Table 1). The two Chinese landraces crossed with Hark (N N) had about 18% [F.sub.1] pollen sterility (Table 1).
[Figure 1 ILLUSTRATION OMITTED]
Table 1. Pollen fertility of [F.sub.1] plants and [F.sub.2] plants from cross-pollinations of homozygous chromosome inversion soybean plants.
[F.sub.1] plants Cross combinations([dagger]) Sterile pollen No. % PI 597651 x PI 597652 2.1 [+ or -] 0.9 5 Hark N N x Hark In 25.2 [+ or -] 2.4 5 In([double dagger]) PI 597651 x Hark N N 17.5 [+ or -] 2.3 5 PI 597652 x Hark N N 18.7 [+ or -] 2.5 5 PI 597651 x Hark In In 48.2 [+ or -] 2.5 5 PI 597652 x Hark In In 46.6 [+ or -] 2.6 5 No. [F.sub.2] plants Sterile pollen Cross combinations([dagger]) 0-10% 11-20% 21-30% no. PI 597651 x PI 597652 20 0 0 Hark N N x Hark In 46 6 46 In([double dagger]) PI 597651 x Hark N N 8 13 4 PI 597652 x Hark N N 8 10 2 PI 597651 x Hark In In 7 6 7 PI 597652 x Hark In In 7 5 7 No. [F.sub.2] plants Sterile pollen Cross combinations([dagger]) 31-40% 41-50% Total PI 597651 x PI 597652 0 0 20 Hark N N x Hark In 6 0 104 In([double dagger]) PI 597651 x Hark N N 0 0 25 PI 597652 x Hark N N 0 0 20 PI 597651 x Hark In In 3 2 25 PI 597652 x Hark In In 3 3 25
([dagger]) N N is homozygous normal chromosome structure, and In In is homozygous inversion chromosome structure.
([double dagger]) In In refers to the backcross of PI 407179 to cultivar Hark to create a near-isogenic homozygous inversion line.
The [F.sub.2] Hark cross-combination (N N x In In) gave 46 plants with very low percentage of pollen sterility and 46 plants with a level of pollen sterility similar to the [F.sub.1] plants (Table 1). A few [F.sub.2] plants had slightly lower or slightly higher pollen sterility. The two Chinese landraces crossed with Hark (N N) gave [F.sub.2] plants that were distributed into three pollen sterility categories (Table 1).
The two Chinese landraces crossed with Hark (In In) gave [F.sub.2] plants that were distributed into four sterility categories (Table 1). These pollen sterility categories would be expected to represent no inversion, the Chinese landrace inversion, the Hark (G. soja) inversion, and the two different inversions. The percentage of pollen sterility is dependent on the amount of crossing over within the inverted region. A pollen sterility class of 31 to 40% would include [F.sub.2] plants with variation in crossover frequencies (Dowrick 1957). Suppression of crossing over by asynapsis and nonhomologous pairing would lower the number of crossovers, thereby reducing the percentage of pollen abortion (McClintock, 1931; Maguire, 1966). A pollen sterility class of 41 to 50% would represent those plants with both paracentric inversions.
The minimum genetic length of an inversion may be estimated from pollen abortion frequencies (Morgan, 1950). As a correction, the percentage of pollen abortion occurring in homozygous plants should be deducted from the percentage of pollen abortion in heterozygous inversion plants. The control [F.sub.1] plants grown in Ames, IA had 1% aborted pollen. Therefore, based on pollen data, the minimum genetic length of this inversion is 25.2% - 1.0% = 24.2%. This calculation considers that single crossovers and the average of two-strand, three-strand, and four-strand doubles will produce tetrads with two normal and two aborted microspores (Burnham, 1962). The two Chinese landraces also had about 1% aborted pollen in control plants. Thus the minimum genetic length for this inversion should be reduced by about 1% and becomes 16.5% for PI 597651 and 17.7% for PI 597652.
The cross-combination Hark (N N) x Hark (In In) was grown in a glasshouse at Ames, IA, and in the field at Gongzhuling, China. In the Iowa environment, bridges plus fragments were observed in 19.4% of the meiocytes (Fig. 1C and 1D, Table 2). This contrasts to only 2.8% meiotic aberrations for the same cross-combination grown in the China environment (Table 2). Differences in growth conditions may be the explanation for the apparent discrepancy in the number of meiotic aberrations between the same genotype grown in two different environments.
Table 2. Meiotic irregularities of [F.sub.1] plants from cross-pollinations of homozygous inversion soybean plants.
Anaphase I No. laggards No. No. and Cross combinations([dagger]) cells bridges fragments Hark N N x Hark In 100 7 13 In([double dagger]) Hark N N x Hark In In 36 0 1 PI 597651 x Hark N N 17 3 3 PI 597652 x Hark N N 60 12 12 PI 597651 x Hark In In 43 5 4 PI 597652 x Hark In In 45 5 10 Telophase I No. laggards No. No. and Cross combinations([dagger]) cells bridges fragments Hark N N x Hark In NE([sections]) - - In([double dagger]) Hark N N x Hark In In 19 0 0 PI 597651 x Hark N N 41 10 6 PI 597652 x Hark N N 34 0 8 PI 597651 x Hark In In 3 0 0 PI 597652 x Hark In In 214 0 8 Telophase I Metaphase II No. laggards No. No. and Cross combinations([dagger]) cells bridges fragments Hark N N x Hark In NE - - In([double dagger]) Hark N N x Hark In In 37 1 2 PI 597651 x Hark N N 46 9 4 PI 597652 x Hark N N 36 8 0 PI 597651 x Hark In In 20 1 0 PI 597652 x Hark In In 12 8 0 Anaphase II No. laggards No. No. and Cross combinations([dagger]) cells bridges fragments Hark N N x Hark In 29 3 2 In([double dagger]) Hark N N x Hark In In 7 0 0 PI 597651 x Hark N N 1 0 0 PI 597652 x Hark N N 1 0 0 PI 597651 x Hark In In 115 27 9 PI 597652 x Hark In In 23 4 1 Telophase II No. laggards Cross combi- No. No. and No. nations([dagger]) cells bridges fragments cells Hark N N x Hark In NE - - 129 In([double dagger]) Hark N N x Hark In In 42 0 0 141 PI 597651 x Hark N N 0 0 0 105 PI 597652 x Hark N N 10 1 0 141 PI 597651 x Hark In In 1 0 0 182 PI 597652 x Hark In In 6 1 1 110 Total Bridges Laggard + laggards Bridges fragments and fragments Cross combi- nations([dagger]) No. % No. % No. % Hark N N x Hark In 10 7.8 15 11.6 25 19.4 In([double dagger]) Hark N N x Hark In In 1 0.7 3 2.1 4 2.8 PI 597651 x Hark N N 22 20.9 13 12.4 35 33.3 PI 597652 x Hark N N 21 14.9 20 14.2 41 29.1 PI 597651 x Hark In In 33 18.1 13 7.1 46 25.2 PI 597652 x Hark In In 18 16.3 20 18.2 38 34.5
[dagger] N N is homozygous normal chromosome structure, and In In is homozygous inversion chromosome structure.
([double dagger]) [F.sub.1] plants grown in glasshouse at Ames, IA, with 24 [+ or -] 2 [degrees] C (dark) and 29 [+ or -] 2 [degrees] C (light), the other five cross-combination [F.sub.1] plants were grown in pots outside at Gongzhuling, China, with 17 [+ or -] 2 [degrees] C (dark) and 22 [+ or -] 2 [degrees] C (light).
([sections]) not examined.
More than 100 meiocytes were examined cytologically for the Hark (N N) x Hark (In In) cross-combination. Different scientists made the observations, but that variable should not account for the sevenfold variation in values. Temperature is known to influence crossing over in plants (Dowrick 1957). Percentage of pollen sterility is dependent on the number of crossovers that occur within inversion loops (Morgan 1950). Ahmad et al. (1977b, 1983) demonstrated conclusively that high temperatures increased meiotic aberrations in soybean inversion heterozygotes. Our data from the Hark (N N) x Hark (In In) plants, grown in two contrasting environments, probably are the result of the effect of temperature differences on meiosis. The higher temperatures in Iowa seemed to increase the frequency of bridges plus laggards and fragments. Our data are consistent with the observations of Ahmad et al. (1977b).
In the cross PI 597651 x Hark (N N), almost twice as many cells with bridges were recorded as cells with laggards and fragments (Fig. 1E, Table 2). However, in the cross PI 597652 x Hark (N N), about equal numbers of cells with bridges were recorded as cells with laggards and fragments (Table 2).
Almost three times as many cells with bridges were recorded as cells with laggards and fragments in the cross PI 597651 x Hark (In In) (Fig. 1F, Table 2). About equal numbers of cells with bridges were recorded as cells with laggards and fragments in the cross PI 597652 x Hark (In In) (Table 2).
The meiotic aberrations observed in the two cross-combinations with PI 567651 resulted in more bridges than laggards and fragments (Table 2). Different configurations of dicentric bridges and fragments at anaphase I and II are expected and depend on the types of crossovers within the inversion loop (Singh, 1993). Crossover configurations that gave rise to dicentric chromatids were consistently higher in cross-combinations with PI 567651 than with PI 567652. In addition, cryptic structural differences in chromosomes (Stephens, 1950) between the two Chinese G. max landraces may have influenced chromosome pairing in the crosses with Hark (N N) and its near-isogenic homozygous inversion line (In In).
(1) The mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA or Iowa State University and does not imply its approval to the exclusion of other products that may be suitable.
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R. G. Palmer,(*) H. Sun, and L. M. Zhao
R.G. Palmer, USDA ARS CICGR Unit and Departments of Agronomy and of Zoology/Genetics, Iowa State University, Ames, Iowa 50011-1010; H. Sun and L.M. Zhao, Jilin Academy of Agricultural Sciences, Gongzhuling, Jilin Province 136100, China. The first two authors contributed equally to this work. This is a joint contribution, Journal Paper J-18412, of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA (project 3352), supported by Hatch Act and State of Iowa, and from the United States Department of Agriculture, Agricultural Research Service, Corn Insects and Crop Genetics Research Unit. Received 28 July 1999. (*) Corresponding author (email@example.com).
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|Author:||Palmer, R. G.; Sun, H.; Zhao, L. M.|
|Date:||May 1, 2000|
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