A. I. Shchapova

[Production of wheat-rye substitution lines and identification of chromosome composition of karyotypes using C-banding, GISH, and SSR markers].

Based on the cross (Triticum aestivum L. × Secale cereale L.) × T. aestivum L., wheat-rye substitution lines (2n = 42) were produced with karyotypes containing, instead of a pair of homologous wheat chromosomes, a homeologous pair of rye chromosomes. The chromosome composition of these lines was described by GISH and C-banding methods, and SSR analysis. The results of genomic in situ hybridization demonstrated that karyotype of these lines included one pair of rye chromosomes each and lacked wheat-rye translocations. C-banding and SSR markers were used to identify rye chromosomes and determine the wheat chromosomes at which the substitution occurred. The lines were designated 1R(1D), 2R(2D)2, 2R(2D)3, 3R(3B), 6R(6A)2. The chromosome composition of lines 1R(1A), 2R(W)1, 5R(W), 5R(5A), and 6R(W)1, which were earlier obtained according to the same scheme for crossing, was characterized using methods of telocentric analysis, GISH, C-banding, and SSR analysis. These lines were identified as 1R(1A), 2R(2D)1, 5R(5D), 5R(5A), and 6R(6A)1, C-banding of chromosomes belonging to line 1R(1A) revealed the presence of two translocated chromosomes (3DS.3DL-del. and 4AL.W) during simultaneous amplification of SSR markers located on 3DL and 4AS arms. The “combined” long arm of the newly derived chromosome 4A is assumed to be formed from the long arm of chromosome 4AS itself and a deleted segment 3DL. All examined lines are cytologically stable, except for 3R(3B), which does not affect the stability of rye 3R chromosome transfer. Chromosome identification and classification of the lines will permit them to be models for genetic studies that can be used thereafter as promising “secondary gene pools” for the purpose of plant breeding.

[Mechanisms of meiotic restitution and their genetic regulation in wheat-rye polyhaploids].

Meiosis has been studied in partially fertile wheat–rye F1 hybrids yielded by crosses Triticum aestivum (Saratovskaya 29 variety) × Secale cereale L. (Onokhoiskaya variety) (4x =28). Hybrid self-fertility proved to be caused by formation of restituted nuclei, which appear after equational segregation of univalent chromosome in AI and sister chromatid non-separation in AII of meiosis, as well as after AI blockage in three different ways. Both types of meiotic restitution were found in each hybrid plant. Expression of the “meiotic restitution” trait varied significantly in polyhaploids of the same genotype (ears of the same plants, anthers of the same ear, microsporocytes of the same anther). Chromatin condensation in prophase proved to be related to the division type and univalent segregation in AI. During reduction segregation of univalents in AI, sister chromatid cohesion and chromosome supercondensation remained unchanged. The results obtained suggest that in the remote hybrids with haploid karyotype of the parental origin (polyhaploids), the program of two-stage meiosis may be fundamentally transformed to ensure one instead of two divisions. We propose that meiotic restitution is a result of special genetic regulation of the kinetochore organization (both structural and functional) and chromatin condensation, i.e. of major meiotic mechanisms.

Meiotic restitution in amphihaploids in the tribe Triticeae

In haploid and diploid organisms of the plant kingdom, meiotic division of diploid cells proceeds in two consecutive stages, with DNA replicating only once. In amphihaploids (interspecific or intergeneric hybrids), where homologs are absent, the reduction of the chromosome number does not occur, meiosis is abnormal, and the plants are sterile. Gamete viability in F1 hybrids is ensured by a single division when chromosomes are separated into sister chromatids in either the first or the second division. Such gametes ensure partial fertility of amphihaploids, thereby facilitating their survival and stabilization of the polygenome. The frequency of the formation of viable gametes varies from a few cases to 98.8% in different anthers of the hybrids. Here, studies on the cytological mechanisms and genetic control of chromosome unreduction or restitution in different amphihaploids of the tribe Triticeae are reviewed. The current notions on the control of formation of restitution nuclei based on the principles of a prolonged metaphase I and different types of meiocytes. The main terms used for systematization of restitution mechanisms are first-division restitution (FDR), single-division meiosis (SDM), and unreductional meiotic cell division (UMCD). It has been assumed that archesporial cells of wide hybrids may have two cell division programs, the meiotic and the mitoyic ones The possible approaches to the analysis of the genetic control of chromosome restitution in amphihaploids are discussed.

Role of rye chromosome 2R from wheat-rye substitution line 2R(2D)1 (Triticum aestivum L. cv. Saratovskaya 29-Secale cereale L. cv. Onokhoiskaya) in genetic regulation of meiotic restitution in wheat-rye polyhaploids

A study was made of the role of rye chromosome 2R from the wheat-rye substitution line 2R(2D)1 (Triticum aestivum L. cv. Saratovskaya 29-Secale cereale L. cv. Onokhoiskaya) in genetic regulation of meiotic restitution in wheat-rye polyhaploids 2R(2D)1 × S. cereale L. cv. Onokhoiskaya. Rye chromosome 2R proved to affect the completeness of the meiotic program, suppressing the formation of restitution gametes. This was evident from the reductional division of univalent chromosomes in AI and the occurrence of the second meiotic division. The interrelationships between the type of chromosome division in AI and the two-step character of meiosis are discussed. The structural and functional organization of the centromeric regions of chromosomes undergoing reductional division is assumed to determine the two-step character of division.

Production of wheat-rye substitution lines based on winter rye cultivars with karyotype identification by means of C-banding, GISH, and SSR markers

The study presents a continuation of the research aimed at producing of wheat-rye substitution lines (2n = 42) based on the cross (Triticum aestivum L. × Secale sereale L.) × Triticum aestivum L., and using winter rye cultivars Vyatka and Vietnamskaya Mestnaya. In BC 1 F 5 two lines were identified, having karyotypes in which a pair of homologous wheat chromosomes was substituted by a homeologous pair of rye chromosomes. The chromosome composition of these lines was analyzed using C-banding, GISH, and SSR markers. It was demonstrated that karyotype of each line included a single pair of rye chromosomes and lacked wheat-rye translocations. The rye chromosomes were identified, and the chromosomes of wheat, at which the substitutions occurred, were determined. The lines generated by crosses with rye of Vyatka and Vietnamskaya Mestnaya cultivars were designated 1Rv(1A) and 5Rviet(5A), respectively. Chromosome identification and classification of the lines makes it possible to use them in breeding programs and genetic studies.

Comparative Effects of Rye Chromosomes 1R and 5R on Androgenesis in Cultured Anthers of Wheat-Rye Substitution Lines as Dependent on the Line Origin

The effects of rye chromosomes 1R and 5R on androgenesis in cultured anthers of wheat–rye substitution lines was studied as dependent on the cultivar origin of the rye chromosomes and on the wheat genome (A or D) subjected to substitution. Chromosome 1R stimulated embryogenesis in anther cultures, while chromosome 5R suppressed it regardless of whether the corresponding wheat chromosomes were substituted in the A or D genome. The effect of chromosome 1R on embryogenesis proved to depend on its cultivar origin. Along with rye chromosome 1R, wheat chromosome 1A was shown to substantially affect total seedling regeneration. Regeneration of green seedlings was dramatically affected both by rye chromosome 1R and by wheat chromosome 1D. The results supported the published data that individual androgenesis parameters (embryogenesis, total plant regeneration, green plant regeneration) are controlled by different genetic mechanisms.

Features of the regulation of meiotic restitution in androgenic haploids of wheat-rye substitution lines 2R(2D)1, 2R(2D)3, and 6R(6A) (Triticum aestivum L., cultivar Saratovskaya 29/Secale cereale L., cultivar Onokhoiskaya)

Regulation of meiotic restitution in androgenic haploids generated by cultivation of isolated anthers of three wheat-rye substitution lines 2R(2D)1, 2R(2D)3, and 6R(6A) (Triticum aestivum L., cultivar Saratovskaya 29/Secale cereale L., cultivar Onokhoiskaya) was studied. The presence of rye chromosomes and the absence of homeologous wheat chromosomes in the haploid plant genome was shown to cause meiotic restitution, as observed in the case of androgenic haploids 6R(6A), or to inhibit it—in meiosis of haploids 2R(2D)1 and 2R(2D)3. In haploids of lines 2R(2D)1 and 2R(2D)3, the reductional type of division of univalent chromosomes was observed, leading to preferential formation of tetrads. In haploids of line 6R(6A), the equational type of division of univalents into sister chromatids, resulting in the block of the second division and formation of diads in approximately 50% of cells, was detected. These results confirm data on the effect of the genotype of line 2R(2D)1 on the induction of reductional type division of univalents and two-phase meiosis, which were earlier obtained in studies of meiosis in polyhaploids 2R(2D)1 × S. cereale L., cultivar Onokhoiskaya.

Genetic regulation of the centromere division in rye and wheat univalent chromosomes in dimonosomics during the meiotic anaphase I

Cytogenetic analysis of meiosis in the wheat-rye dimonosomics 1Rv-1A, 1Ron-1A, 2R-2D, 5R-5A, and 6R-6A was conducted. C-banding was used to study the segregation pattern of each of two univalent chromosomes during the first meiotic division. It has been shown that the division frequency of the centromeric regions of all rye chromosomes in the pair studied is significantly higher than in the wheat chromosomes. The ANOVA performed suggest that the plant genotype contributes significantly (at P = 0.05) to the behavior pattern of univalent chromosomes in meiosis. The data obtained demonstrate that the rye and wheat chromosomes studied are involved in genetic regulation of centromere division in meiotic anaphase I (AI). The presence of rye chromosome 2R and wheat chromosome 2D suppresses the division of centromeres of the sister chromatids in AI. Rye chromosomes 1Rv, 1Ron, 5R, and 6R induce equational division; however, rye chromosome 1Rv increases to a greater degree the frequency of equational division of wheat chromosome 1A as compared with chromosome 1Ron.

Effect of Rye Secale cereale L. Chromosomes 1R and 3R on Polyembryony Expression in Hybrid Combinations between ( Hordeum L.)-Triticum aestivum L. Alloplasmic Recombinant Lines and Wheat-Rye Substitution Lines T. aestivum L.- S. cereale L.

The effect of rye chromosomes on polyembryony was studied for reciprocal hybrid combinations between (Hordeum vulgare L.)-Triticum aestivum L. alloplasmic recombinant lines and five wheat T. aestivum L. (cultivar Saratovskaya 29)-rye Secale cereale L. (cultivar Onokhosikaya) substitution lines: 1R(1D), 2R(2D), 3R(3B), 5R(5A), and 6R(6A), and for direct hybrid combinations between the [H. marinum ssp. gussoneanum (H. geniculatum All.)]-T. aestivum alloplasmic recombinant line and the wheat-rye substitution lines 1R(1A), 1R(1D), and 3R(3B). Chromosomes 1R and 3R of rye cultivar Onokhoiskaya proved to affect the expression of polyembryony in the hybrid combinations that involved the alloplasmic recombinant lines of common wheat as maternal genotypes. Based on this finding, polyembryony was regarded as a phenotypic expression of nuclear-cytoplasmic interactions where an important role is played by rye chromosomes 1R and 3R and the H. vulgare cytoplasm. Consideration is given to the association between the effect of rye chromosomes 1R and 3R on polyembryony in the [(Hordeum)-T. aestivum × wheat-rye substitution lines] hybrid combinations and their stimulating effect on the development on androgenic embryoids in isolated anther cultures of the wheat-rye substitution lines.

The effect of rye chromosomes on the characteristic features of androgenesis in wheat-rye substitution lines Triticum aestivum L. (cv. Saratovskaya 29)/Secale cereale L. (cv. Onokhoiskaya) and triticale

The characteristic features of androgenesis in six wheat–rye substitution lines Triticum aestivumL. (cv. Saratovskaya 29)/Secale cerealeL. (cv. Onokhoiskaya) and triticale (2n= 56) using anther culture at different concentrations of 2,4-D in the growth medium were studied. Under variable cultivation conditions, the significant effect of genotypic diversity on the variability of such androgenesis parameters as the frequency of productive anthers, the frequency of embryoid formation, and the frequency of total regenerated plantlets, was shown. It was demonstrated that chromosomes 1R, 3R, and 7R stimulated the formation of androgenous embryoids, while chromosome 5R produced an opposite effect. In triticale and substitution lines, the regeneration ability of androgenous embryoids induced by elevated 2.4-D concentrations was inhibited. Chromosome 1R of the Onokhoiskaya cultivar was suggested to contain genes suppressing regeneration of green plantlets, while chromosome 3R, conversely, stimulated their formation. Chromosomes 1R, 2R, 3R, and 7R of the Onokhoiskaya cultivar did not inhibit the spontaneous formation of androgenous hexaploids in the substitution lines.