Genetic and cytological analyses of a partial-female-sterile mutant (PS-1) in soybean (Glycine max; Leguminosae)

Soybean partial-female-sterile mutant 1 (PS-1) was recovered from a gene-tagging study. The objectives were to study the inheritance, linkage, allelism, and certain aspects of the reproductive biology of the PS-1 mutant. For inheritance and linkage tests, PS-1 was crossed to flower color mutant Harosoy-w4 and to chlorophyll-deficient mutant CD-1, also recovered from the gene-tagging study. For allelism tests, reciprocal crosses were made with PS-1 and three other partial-sterile mutants (PS-2, PS-3, and PS-4) recovered from the same gene-tagging study. The PS-1 mutant is inherited in a 3:1 ratio and is a single recessive gene. Linkage results indicated that the gene for partial sterility in PS-1 is not linked either to the w4 locus or to the CD-1 locus. Allelism tests showed that the gene in PS-1 is nonallelic to the gene in PS-2, PS-3, and PS-4. Investigations of developing and mature pollen indicated no differences in morphology, stainability, or fluorescence between normal and partial-sterile genotypes. The PS-1 mutant is completely male fertile. Confocal scanning laser microscopy was used to determine that early embryo abortion in PS-1 is due indirectly to abnormal migration of the fused polar nucleus, which prevented it from being fertilized. Subsequent absence of endosperm development leads directly to abortion of the proembryo.     

Genetics and cytology of a genic male-sterile, female-sterile mutant from a transposon-containing soybean population

A male-sterile, female-sterile soybean mutant (w4-m sterile) was identified among progeny of germinal revertants of a gene-tagging study. Our objectives were to determine the genetics (inheritance, allelism, and linkage) and the cytology (microsporogenesis and microgametogenesis) of the w4-m sterile. The mutant was inherited as a single recessive nuclear gene and was nonallelic to known male-sterile, female-sterile mutants st2 st2, st3 st3, st4 st4, st5 st5, and st6 st6 st7 st7. No linkage was detected between the w4-m sterile and the w4w4, y10 y10, y11 y11, y20 y20, fr1 fr1, and fr2 fr2 mutants. Homologous chromosome pairing was complete in fertile plants. Chromosome pairing, as observed in squash preparation, was almost completely absent in sterile plants. Developmentally microsporogenesis proceeded normally in both the fertile and the w4-m sterile through the early microspore stage. Then the tapetal cells of the w4-m sterile surrounding the young microspores developed different-size vacuoles. These tapetal cells became smaller in size and separated from each other. Some of the microspores of the w4-m sterile also became more vacuolate prematurely and sometimes they collapsed, usually by the late microspore stage. In the w4-m sterile the microspore walls remained thinner and structurally different from the microspore walls of fertile plants. No pollen was formed in the mutant plants, even though some of the male cells reached the pollen stage, although without normal filling. The w4-m sterile was designated st8st8 and assigned Soybean Genetic Type Collection number T352.     

Genetic analyses of two independent chlorophyll-deficient mutants identified among the progeny of a single chimeric foliage soybean plant

Chimeric (variegated) foliage plants are frequently observed in many species. In soybean [Glycine max(L.) Merr.], progeny of chimeric plants are a source of nuclear and cytoplasmically inherited mutants. Self-pollinated progeny of a single chimeric plant derived from tissue culture of PI 427099 (Jilin 3) included plants with green foliage, chimeric foliage, yellow foliage (viable), and yellow foliage (lethal). Our objectives were to determine (1) inheritance, linkage, and allelism of the lethal-yellow mutant with known chlorophyll-deficient mutants; (2) inheritance, linkage, and allelism of the viable-yellow mutant with known chlorophyll-deficient mutants; (3) allelism of the lethal-yellow mutant with the viable-yellow mutant; and (4) male and female gamete transmission of the viable-yellow mutant trait. The viable-yellow mutant was allelic to T323, y20 y20 (Ames 2) Mdh1-n Mdh1-n (Ames 2) and was assigned genetic type collection number T361 and gene symbol y20 y20 (Ames 24) Mdh1-n Mdh1-n (Ames 22). The lethal-yellow mutant was allelic to T225H (Y18 y18) and was assigned genetic type collection number T362H and gene symbol Y18 y18 (Ames 2). T225H became Y18 y18 (Ames 1). The two chlorophyll-deficient mutants were not linked to each other. There was no significant difference in F(1) male or female gamete transmission of the viable-yellow mutant. However, many cross-combinations gave significant deviations from the expected 3 green plants:1 viable-yellow plant in the F(2) generation. The allelism of these two chlorophyll-deficient mutants with mutants T225H and T323, derived from putative transposable element systems, is intriguing. An explanation of this phenomenon awaits molecular experimentation.     

Duplicate chlorophyll-deficient loci in soybean.

Three lethal-yellow mutants have been identified in soybean (Glycine max (L.) Merr.), and assigned genetic type collection numbers T218H, T225H, and T362H. Previous genetic evaluation of T362H indicated allelism with T218H and T225H and duplicate-factor inheritance. Our objectives were to confirm the inheritance and allelism of T218H and T225H and to molecularly map the locus and (or) loci conditioning the lethal-yellow phenotype. The inheritance of T218H and T225H was 3 green : 1 lethal yellow in their original parental source germplasm of Glycine max 'Illini' and Glycine max 'Lincoln', respectively. In crosses to unrelated germplasm, a 15 green : 1 lethal yellow was observed. Allelism tests indicated that T218H and T225H were allelic. The molecular mapping population was Glycine max 'Minsoy' x T225H and simple sequence repeat (SSR) markers were used. The first locus, designated y18-1, was located on soybean molecular linkage group B2, between SSR markers Satt474 and Satt534, and linked to each by 4.4 and 13.4 cM, respectively. The second locus, designated y18-2, was located on soybean molecular linkage group D2, between SSR markers Satt543 and Sat-001, and linked to each by 2.2 and 4.4 cM, respectively.     

Inheritance of malate dehydrogenase nulls in soybean

Three chlorophyll-deficient mutants (CD-1, CD-2, and CD-3), derived from the progeny of independent germinal revertants from the w4-mutable soybean line [Glycine max (L.) Merrill], were characterized genetically. Electrophoretic analyses indicated that these lines lacked two of three mitochondrial malate dehydrogenase isozymes (MDH-). The absence of two MDH bands was conditioned by a recessive allele at a locus designated Mdh1. All three CDs were allelic to each other and to T253, a Harosoy isoline y20-k2 MDH- from the Genetic Type Collection. The MDH- phenotype and the yellow-green plant phenotype were each inherited as single recessive alleles. No recombination between the two traits was found in nine F2 populations from crosses of the CDs by wild-type soybean lines. Complete linkage of the Mdh1 and y20 loci suggested that the mutations in the chlorophyll-deficient lines were deletions. Phenotypic differences among the CDs suggested that the deletions may have different endpoints. The chromosomal aberrations were not large enough to affect transmission of y20 and Mdh1 mutant alleles through the pollen or ovule. CD-1, CD-2, and CD-3 were added to the Soybean Genetic Type Collection as T323, T324, and T325, respectively.     

Molecular mapping of four ovule lethal mutants in soybean

We report genetic mapping of four soybean ovule lethal mutants, PS-1, PS-2, PS-3, and PS-4, which had been identified as female partial-sterile mutants from a gene-tagging study. The four mutants had been classified into two mutation classes: (1) PS-1—sporophytic mutation affects sporophytically expressed genes; and (2) PS-2, PS-3, and PS-4 mutants—female gametophyte-specific mutations affect gametophytically expressed genes and are transmitted through the male, but not the female gametes. Molecular mapping demonstrated that these four mutant genes and previously reported female-partial sterile gene, Fsp1, are located independently on soybean molecular linkage groups (MLG-) using SSR markers. PS-1, designated as Fsp2 and Genetic Type Collection number T364, is located between SSR markers Satt170 and Satt363 on MLG-C2 and linked by 13.9 cM and 12.1 cM, respectively. PS-2, designated as Fsp3 and Genetic Type Collection number T365H, is located between SSR markers Satt538 and Satt429 on MLG-A2 and linked by 13.3 cM and 25.4 cM, respectively. PS-3, designated as Fsp4 and Genetic Type Collection number T366H, is located on the terminus of MLG-F and linked to Sat 152 by 13.1 cM. PS-4, designated as Fsp5 and Genetic Type Collection number T367H, is located between SSR markers Satt324 and Satt138 on MLG-G and linked by 19.6 cM and 7.5 cM, respectively. SSR markers adjacent to Fsp3, Fsp4, and Fsp5 were distorted from a 1:2:1 ratio and fit a 1:1 ratio. The segregation distortions of SSR markers adjacent to Fsp3, Fsp4, and Fsp5 are in support of male, but not female transmission of the Fsp3, Fsp4, and Fsp5 gametes.This is a joint contribution of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, Project No. 3769 and from the USDA, Agricultural Research Service, Corn Insects and Crop Genetics Research Unit, and supported by Hatch Act and State of Iowa. The mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by Iowa State University or the USDA, and the use of the name by Iowa State University or the USDA implies no approval of the product to the exclusion of others that may also be suitable.Communicated by J. Dvorak     

mRNA differential display between sterile and fertile anther of rice and analysis of cDNA differential fragments

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Investigating the genetic model for brown stem rot resistance in soybean

Genetic analyses have indicated that brown stem rot (BSR) resistance in soybean is conferred by dominant alleles at three independent loci, the actions of which may be modified by linked or independent loci. A study was conducted to characterize the inheritance of BSR resistance in PI 567609, a soybean plant introduction from China. Segregating progeny from crosses of PI 567609 with BSR-susceptible and -resistant genotypes were evaluated for response to BSR-causal fungus, Phialophora gregata. Genetic analyses indicated that PI 567609 carries a single gene or cluster of linked genes for brown stem rot resistance, and that this gene (or cluster) is allelic to, or tightly linked to previously identified resistance genes, Rbs1, Rbs2, and Rbs3. Because previous allelism tests indicated that Rbs1, Rbs2, and Rbs3 were unlinked, and molecular mapping studies have indicated that Rbs1, Rbs2, and Rbs3 are linked on molecular linkage group J of soybean, a new model is proposed for BSR resistance. In this model, BSR resistance is controlled through the interaction of alleles at four independent loci, at least two of which are necessary to condition a resistance response. Functional redundancy at three of these loci allows any one of the three to interact with a fourth locus to confer resistance to BSR.     

Allelism tests of 15 dominant cataract mutations in mice.

Fifteen autosomal dominant mutations that cause cataract of lenses in mice were tested for allelism. The outcrosses of double mutants revealed three allelism groups, consisting of 5, 4 and 2 mutations as well as 4 mutations which segregated independently. The results indicated 7 different cataract loci in the sample of 15 mutations. The biomicroscopic examination of the eyes showed that phenotypically similar as well as very distinct cataract mutations can be alleles of the same gene. Conversely, phenotypically similar mutations were shown to be non-allelic.     

The timing of mu activity in maize

The timing of mutator activity of Mu in maize was tested in three ways: (1) by allelism tests of phenotypically similar male-transmitted mutants, (2) by studying the clustering of phenotypically similar mutants as demonstrated by ear maps and the subsequent allelism tests of these mutants, and (3) by the induction of somatic sectors in Mu plants heterozygous for plant and endosperm marker genes. Allelism tests of phenotypically similar mutants in outcrosses of Mu plants as males established that 18.6% were allelic and that premeiotic mutants are induced. This conclusion was supported by ear maps of Mu-bearing plants, which revealed sectors of seeds that produced plants bearing phenotypically similar allelic mutants. The smallness of these sectors indicated that the premeiotic activity of Mu that gave rise to them occurred very late. The lack of visible sectors in mature sporophytic, endosperm and aleurone tissue in plants carrying Mu supports the conclusion that the mutator activity of Mu does not occur throughout the ontogeny of the plant and seems to be restricted to a time shortly before and/or during meiosis.     

The isolation, partial characterization, and biosynthesis of the paragonial substances, PS-1 and PS-2 of Drosophila funebris

In methanolic extracts of accessory glands (paragonia) from Drosophila funebris, two specific, ninhydrin-positive substances, PS-1 and PS-2, were found. PS-1 and PS-2 were isolated by column chromatography. PS-1 consists of 27 amino acid residues. Two forms of PS-1 are present in the ratio of 7:3 which differ only in the content of valine and leucine. Fractions containing partially purified valine-PS-1 and leucine-PS-1, respectively, have the same biological activity. All males in the population synthesize both forms of PS-1. PS-2 is a low molecular weight substance containing glycine and ammonia as ninhydrin-positive components and carbohydrate as indicated by several sugar tests. In vitro studies showed that copulation provides the stimulus for enhanced synthesis of paragonial substances.     

Properties of mutant acetolactate synthases resistant to triazolopyrimidine sulfonanilide

Triazolopyrimidine sulfanilides are a class of highly active herbicides whose primary target is acetolactate synthase. Spontaneous mutants of tobacco (Nicotiana tabacum) (KS-43) and cotton (Gossypium hirsutum) (PS-3 and DO-2) resistant to triazolopyrimidine sulfonanilide were selected in tissue culture. Acetolactate synthase partially purified from the three mutants were 80- to 1000-fold less sensitive to inhibition by the compound compared with the corresponding wild-type enzyme. The mutants also varied in the cross-resistance pattern to other acetolactate synthase inhibiting herbicides in the sulfonylurea, imidazolinone, and pyrimidyl-oxy-benzoate chemical families. Thus, acetolactate synthase from KS-43, PS-3, and DO-2 cultures have different mutations. The affinities for pyruvate, thiamine pyrophosphate, as well as the activity of the mutant enzymes were found to be comparable to the corresponding wild-type enzymes. However, the enzyme from PS-3 was highly resistant to feedback inhibition by valine and leucine. In contrast, acetolactate synthase from KS-43 and DO-2 were inhibited by valine and leucine to nearly the same extent as the wild-type enzymes. Also, PS-3 cultures accumulated much higher levels of the branched chain amino acids compared to the wild-type cotton culture. The mutation in the PS-3 enzyme has therefore rendered it insensitive to feedback regulation by valine and leucine.     

Localization and fine mapping of gaMS-1, a male gametophytic mutant of maize

Post-meiotic mutants affecting pollen development are fundamental tools for defining the genetic program controlling microsporogenesis and pollen function. An example of such mutants is gametophytic male sterile-1 (gaMS-1). Heterozygous plants gaMS-1/+ that have a normal phenotype and are female fertile, segregate 1:1 normal:sterile pollen grains and their selfed progeny segregates 1:1 normal:semi-sterile plants. With the final aim of isolating the gene, a positional cloning strategy was adopted. In this paper, we report the results of fine mapping GaMS-1 by different types of molecular markers. Two back crosses were used as mapping populations. They were obtained by crossing the line carrying the mutation with the inbred lines Mo17 and WF9, used as recurrent male parents. Linkage disequilibrium analysis allowed assigning GaMS-1 to the short arm of chromosome 2.By the combined use of SSR, AFLP, PCR markers and ESTs a region of 1 cM containing GaMS-1 was delimited. Received: 15 November 2000 / Revision accepted: 24 May 2001     

Mitochondrial heredity of resistance to 3-(3,4-dichlorophenyl)-1,1-dimethylurea, an inhibitor of cytochrome b oxidation, in Saccharomyces cerevisiae.

3-(3,4-dichlorophenyl)-1,1-dimethylurea (diuron), an inhibitor of cytochrome b oxidation, has been used for the selection of three resistant mutants (diur) of Saccharomyces cerevisiae. The mutant diur-64 exhibits in vivo cross-resistance to antimycin A while diur-34 and diur-1 are more sensitive to antimycin A than the parental strain. The three mutants exhibit mitochondrial inheritance according to the following criteria: mitotic segregation of diuron-resistant and diuron-sensitive diploids is obtained among the diploid progeny of a cross between diur and dius; non-Mendelian segregation of diuron resistance (4:0) is observed in spores of tetrads issued from diuron-resistant diploid; extensive ethidium bromide treatment leads to the formation of Q- mutants which no longer transmit diur and dius alleles. Evidence for two distinct diuron-resistant loci were obtained by allelism tests. Recombination analysis shows that diuron-resistance is not located in the polar region of the mitochondrial genome. The diur loci are not linked to the erythromycin locus since the upper limit in recombinants frequency (26%) for a non-polar region is obtained between diur and eryr. A low recombinants frequency (3%) is observed in crosses between diur-34 mutation and the two mutants cob1 and cob2 suggesting that diur-34 might be located between these two cytochrome-b-deficient loci. The resistance to diuron is also expressed in vitro since the oxidation rates of succinate by sonicated submitochondrial particles from the mutants are clearly less sensitive to diuron than that of the wild type.     

Cytokinin receptors in sporophytes are essential for male and female functions in Arabidopsis thaliana

Arabidopsis has three cytokinin receptors genes: CRE1, AHK2 and AHK3. Availability of plants that are homozygous mutant for these three genes indicates that cytokinin receptors in the haploid cells are dispensable for the development of male and female gametophytes. The triple mutants form a few flowers but never set seed, indicating that reproductive growth is impaired. We investigated which reproductive processes are affected in the triple mutants. Anthers of mutant plants contained fewer pollen grains and did not dehisce. Pollen in the anthers completed the formation of the one vegetative nucleus and the two sperm nuclei, as seen in wild type. The majority of the ovules were abnormal: 78% lacked the embryo sac, 10% carried a female gametophyte that terminated its development before completing three rounds of nuclear division, and about 12% completed three rounds of nuclear division but the gametophytes were smaller than those of the wild type. Reciprocal crosses between the wild type and the triple mutants indicated that pollen from mutant plants did not germinate on wild-type stigmas, and wild-type pollen did not germinate on mutant stigmas. These results suggest that cytokinin receptors in the sporophyte are indispensable for anther dehiscence, pollen maturation, induction of pollen germination by the stigma and female gametophyte formation and maturation.Key words: cytokinin, cytokinin receptor, female gametophyte, male gametophyte, stigma     

The causes of genetic male sterility in 3 soybaen lines

Summary The cause of male sterility in 3 soybean lines, TGM 103-1, N-69-2774 and TGM 242-4 was studied. In TGM 103-1, which was both male and female sterile, two different abnormalities were associated with sterility. Precocious movement of a few chromosomes at the metaphase I stage resulted into the production of non-functional pollen while cells which underwent apparent normal meiotic division had disintergration of the tapetal cell wall immediately after the free microspore stage leading to the starvation and subsequent death of the developing microspores. In lines N-69-2774 and TGM 242-4, both of which were partially sterile, male sterility resulted from a failure of cytokinesis after the telophase II stage. Meiosis proceeded normally but the 4 microspores after telophase II failed to separate into pollen grains and degenerated thereafter.     

Searching for tagged male-sterile mutants of arabidopsis

Although many male-sterile mutants have been identified inArbidopsis thaliana, few of the corresponding genes have been cloned. In order to facilitate cloning of a male sterility gene, 23 of Feldmann's T-DNA-generated, reduced-fertility lines were screened to identify a tagged male-sterile mutation. Malesterile mutants were identified, as well as mutants that were both male and female sterile. Segregation of the kanamycin marker gene in the progeny of 15 of these lines was studied. Forty percent had functional T-DNAs (encoding resistance to kanamycin) inserted at a single locus, the remainder segregating for two or more functional T-DNA inserts. Linkage between T-DNA inserts and mutant phenotype was tested for six lines. In three of these lines, mutations were not linked to a T-DNA insert. In three lines, the mutation segregated with a T-DNA insert.     

Gametophytic expression of genes controlling endosperm development in maize

Summary An indirect approach was adopted to select viable mutants affecting the male gametophytic generation in maize. This approach consists of a selection of endosperm defective mutants followed by a test for gametophytic gene expression, based on the distortion from mendelian segregation and on the measurement of pollen size and pollen sterility. The material used consisted of 34 endosperm defective viable mutants introgressed in B37 genetic background. Complementation tests indicated that the mutation in the collection of mutants affected different genes controlling endosperm development. The study of the segregation in F2 revealed four classes of de (defective endosperm) mutants: (1) mutants in which the mutation does not affect either gametophytic development or function; (2) mutants in which the effect on the gametophyte affects pollen development processes; (3) mutants showing effects on both pollen development and function, and (4) mutants where only pollen tube growth rate is affected. Positive and negative interactions between pollen and style were detected by means of mixed pollination (pollen produced by de/de plants and pollen from an inbred line used as a standard and carrying genes for colored aleurone), on de/de and de/ + plants. Positive interactions were interpreted as methabolic complementation between defective pollen and normal styles.