Meiotic and Isozymic Analyses of Tall Fescue × Giant Fescue Hybrids and Amphiploids1

The meiotic behavior of three tall fescue (Festuca arundinacea, 2n = 6x = 42) genotypes, giant fescue (F. gigantea, 2n = 6x = 42), and their reciprocal F₁ hybrids and C₁, amphiploids was evaluated to determine the parental genomic relationships. Isozyme banding patterns were used to confirm the parental identity of the hybrids and amphiploids. At meta-phase I, the parents had predominantly bivalent pairing. The hybrids had an average of 9.51 I, 16.02 II, 0.12 III, 0.02 IV, and the amphiploids had 2.17 I, 38.82 II, 0.60 III, 0.58 IV, 0.01 V—VIII. The prevalence of bivalent pairing in both hybrids and amphiploids suggested a homoeologous relationship between the six genomes, with four of the six being more closely related. Bivalent pairing in the amphiploids indicated genetic regulation of chromosome pairing. Zymograms were obtained for acid phosphatase (ACPH), alcohol dehydrogenase (ADH), glutamate oxaloacetate transaminase (GOT), malate dehydrogenase (MDH), 6-phosphogluconate dehydrogenase (6-PGD) and phosphoglucoisomerase (PGI). The three tall fescue and giant fescue parents had different zymograms for ACPH, MDH, 6-PGD and PGI; thus, the tall fescue parents of the hybrids and amphiploids could be determined based on the banding patterns of these four enzymes. Phenotypes were determined for ACPH-1, PGI-2 and 6-PGD-1. ACPH-1 may be used to follow the introgression of giant fescue chromatin into a certain tall fescue genotype.     

Molecular cytogenetic identification of wheat-Elymus tsukushiense introgression lines

Elymus tsukushiense Honda (syn. Roegneria kamoji C. Koch) (2n = 6x = 42, S^tsS^tsH^tsH^tsY^tsY^ts) is a hexaploid species, distantly related to bread wheat Triticum aestivum L. em Thell (2n = 6x = 42, AABBDD). Apart from the delineation of evolutionary relationships, this species is a potential source of resistance to scab, a devastating disease of wheat caused by Fusarium graminearum Schw. A standard C-banded karyotype was established identifying all 21 chromosome pairs of E. tsukushiense. By using C-banding and genomic in situ hybridization analyses, three wheat-E. tsukushiense chromosome addition lines, one ditelosomic addition line, and one disomic substitution line were identified in BC₂ progenies from wheat × E. tsukushiense hybrids. Twenty DNA markers specific for the seven homoeologous groups of the Triticeae were used to determine the homoeology of the added E. tsukushiense chromosomes. The E. tsukushiense chromosomes in the addition lines NAU702, NAU703, and NAU701 were identified as belonging to homoeologous groups 1, 3, and 5, and thus, were designated as 1Ets#1, 3Ets#1, and 5Ets#1, respectively. NAU751 was identified as a disomic substitution line with chromosome 3A of wheat replaced by chromosome 3Ets#1. Line NAU702 has a high level of resistance to scab and will be used in chromosomal engineering and development of improved wheat germplasm for scab resistance breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

Interspecific hybrid of Helianthus annuus × H. simulans: Characterization and utilization in improvement of cultivated sunflower (H. annuus L.)

The first success at interspecific hybridization between cultivated sunflower(H. annuus) and a diploid perennial species, H. simulans is reported. The F₁s from both direct and reciprocal directions exhibited dominance of the wild species phenotype and were pollen sterile. Meiosis was irregular in the F₁ hybrids and both univalents and multivalents were observed. Multiplication of the interspecific hybrids was achieved through in vitro culture of nodal sections on Murashige & Skoog medium supplemented with 0.5 mg l^-1 benzyladenine. Fertility of the interspecific hybrids was improved by subjecting the in vitro proliferating shoots to 0.001% colchicine incorporated in the shoot multiplication medium. The amphiploids serve as fertile bridges and facilitate interspecific gene transfer through conventional breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

The influence of Elymus sibiricus L. genome on the diploidization system of wheat

Summary The meiotic behaviour of five tetraploid wheat strains x Elytricum fertile (2n=42 chromosomes, AABBD(SH) genomes) F₁ hybrids has been analysed. Multivalent associations were observed in the hybrids which could be attributed to Elymus sibiricus L. gene (s) somewhat suppressing the activity of the wheat homoelogous pairing control system. This interaction depends on the wheat genotype. The effect was particularly notable when Triticum turgidum var. salomonis was the wheat parent. The possibility of gene transfer from Elymus to wheat is discussed.     

Molecular cytogenetic characterization of Thinopyrum genomes conferring perennial growth habit in wheat- Thinopyrum amphiploids

Seven wheat‐Thinopyrum amphiploids, AT 3425, AgCs, PI 550710, PI 550711, PI 550712, PI 550713 and PI 550714, were evaluated for perennial growth habit in the field. Three of them, AgCs, AT 3425, and PI 550713, were identified as perennials. Fluorescent genomic in situ hybridization (FGISH) patterns of mitotic chromosomes indicated that AgCs had seven pairs of Thinopyrum chromosomes and 21 pairs of wheat chromosomes. PI 550713 and AT 3425 showed similar FGISH patterns of mitotic chromosomes with three pairs of wheat‐Thinopyrum translocated chromosomes, seven pairs of Thinopyrum chromosomes, and 18 pairs of wheat chromosomes. Thinopyrum chromosome pairing in the Fi hybrid of AT 3425 with AgCs demonstrated differences between Thinopyrum genomes in these two amphiploids. Based on chromosome constitutions, pairing and reported pedigrees, AgCs and AT 3425 were identified as a wheat‐Thinopyrum elongatum amphiploid and partial wheat‐Thinopyrum ponticum amphiploid, respectively. Chromosome pairing in the F₁ hybrid between AT 3425 and PI 550713 revealed that these two amphiploids contained the same Thinopyrum genome. Two different Thinopyrum genomes conferring perennial growth habit were identified from the perennial amphiploids and characterized cytogenetically.     

Hybridizing <Emphasis Type="Italic">Brassica rapa</Emphasis> with wild crucifers <Emphasis Type="Italic">Diplotaxis erucoides</Emphasis> and <Emphasis Type="Italic">Brassica maurorum</Emphasis>

Two wide hybrids, Diplotaxis erucoides (2n = 14) × Brassica rapa (2n = 20) and B. maurorum (2n = 20) × B. rapa, were developed using the sequential ovary–ovule culture. Reciprocal crosses failed, possibly as a consequence of strong unilateral incompatibility. The F ₁ hybrids in each combination were completely male sterile and morphologically intermediate to the respective parents. DNA marker polymorphism and chromosome counts confirmed their hybrid nature. High frequency of bivalents in the F ₁ and the presence of trivalents/quadrivalents in the derived amphiploids suggested genomic duplications and homoeology of the parental genomes. Up to three homoeologous pairs between the D. erucoides (D^eD^e) and B. rapa (AA) genomes, and one between B. maurorum (B^mB^m) and B. rapa genomes were observed. Successful synthesis of the F ₁ hybrids and amphiploids of B. rapa with D. erucoides and B. maurorum, and allosyndetic chromosome pairing are expected to permit introgressions of desirable loci into the cultivated Brassica germplasm, especially for resistance to Alternaria brassicae and Albugo candida.     

A New Intergeneric Hybrid between Triticum aestivum L. and Agropyron fragile (Roth) Candargy: Variation in A. fragile for Suppression of the Wheat Ph-Locus Activity

New intergeneric hybrids were obtained between Triticum aestivum L. cv. Tukuho’ (2n = 6x = 42, AABBDD) and Agropyron fragile (Roth) Candargy PGR 8097 (2n = 4x = 28, PPPP) at a frequency of 1.06 %, through the use of direct embryo culture and in ovulo embryo culture. Such hybrids could be used to transfer barley yellow dwarf virus (BYDV) resistance and winterhardiness into bread wheat. The somatic chromosome number in all the hybrid plants was 2n = 5x = 35, as expected. Considerable variation in chromosome pairing was observed among the different hybrid plants. Average meiotic chromosome configuration at metaphase I was 17.29 Is + 6.57 rod Us + 1.97 ring Us + 0.18 III + 0.03 IV + 0.002 VI. The high level of chromosome pairing in some F₁ hybrids was attributed to Ph-suppressor gene(s) present in A. fragile. The hybrids could not be backcrossed to wheat, but amphiploid seeds have been obtained by colchicine treatment.     

Hybrids between durum wheat and Thinopyrum junceiforme: Prospects for breeding for scab resistance

Tetraploid wheatgrass, Thinopyrum junceiforme(2n = 4x = 28; J₁J₁J₂J₂), a wild relative of wheat, is an excellent source of resistance to Fusarium head blight. Intergeneric F₁ hybrids (2n = 4x = 28; ABJ₁J₂) between durum wheat (Triticum turgidum; 2n = 4x = 28; AABB) cultivars Lloyd or Langdon and Th. junceiforme were synthesized. Most of the pairing in F₁ hybrids was between the J₁- and J₂-genome chromosomes. Some pairing occurred between wheat chromosomes and alien chromosomes, resulting in segmental exchange that was confirmed by fluorescent in situ hybridization (FISH). The F₁hybrids were largely male-sterile and were backcrossed, as the female parent, to the respective durum cultivar. Backcrosses from Lloyd × Th. junceiforme hybrids yielded fertile partial amphiploids (2n = 6x = 42; AABBJ₁J₂) as a result of functioning of unreduced female gametes of the hybrid. Lloyd proved to be a more useful durum parent than Langdon in crosses with Th. junceiforme designed to transfer scab resistance genes. Pairing in the amphiploids was characterized by preferential pairing,which resulted in bivalent formation. However, some intergeneric pairing also occurred. Several fertile hybrid derivatives were produced by further backcrossing and selfing. The introduction of alien chromatin into the durum complement was confirmed by FISH. Hybrid derivative lines had significantly lower mean infection scores (p = 0.01), the best showing 10.93% infection, whereas the parental durum cultivars had 70.34% to 89.46% infection. Hybridization with wild relatives may offer an excellent means of introducing scab resistance into durum wheat. This revised version was published online in July 2006 with corrections to the Cover Date.     

Plant Regeneration and Chromosomal Stability in Tissue Cultures of the Hybrids of Elymus canadensis with Psathyrostachys juncea and Secale cereale

Callus was induced from segments of immature inflorescences of Elymus canadensis×Psathyrostachys juncea and Elymus canadensis×Secale cereale F₁ hybrids on MS medium supplemented with 2 mg/l of 2,4-D and plants were regenerated from the calluses on a hormone free MS medium. The plants regenerated from both the hybrids exhibited a high degree of stability in morphology, chromosome number and chromosome pairing. The reasons for this are discussed.     

The inheritance of resistance to powdery mildew in interspecific hybrids and induced amphiploids of Zinnia elegans Jacq. and Z. angustifolia HBK

Summary Sterile interspecific hybrids and colchicine-induced amphiploids of Zinnia elegans Jacq. and Z. angustifolia HBK were examined to determine the mode of inheritance of resistance to Erysiphe cichoracearum DC ex Merat. Fertility was restored through colchicine treatment of two sterile hybrids of species reciprocal parentage which differed in ray petal response to the pathogen. Derived amphiploids were subsequently intercrossed to overcome the lack of segregation for this trait due to genetic control of pairing upon chromosome doubling. Resistance to E. cichoracearum appears to be complexly inherited in both leaves and ray florets of sterile hybrids and induced amphiploids. Two major dominant genes have been implicated in conferring resistance in ray petal tissue of derived amphiploids. Data obtained from the F₁ hybrid progeny of the intercrossed amphiploids indicate that this trait is not cytoplasmically inherited. It is speculated that the genes conferring resistance in the ray florets are acting independently from those controlling leaf resistance and that most, if not all, of the resistance genes are inherited from Z. angustifolia.Scientific Article No. A-3825, Contribution No. 6804 of the Maryland Agricultural Experiment Station.     

The Tolerance to Aluminum of Triticum N-Genome Amphiploids

Aluminum limits wheat (Triticum aestivum L.) yields on acid soils. An aluminum-tolerant, N-genome Triticum species was used to produce amphiploids, which were tested lor tolerance to 0.44 mM aluminum in solution, to assess the possibility of transferring tolerance to bread wheat. Two types of amphiploids, having the N-genomc (Triticum uniaristatum) in common, were produced by treating three different Triticum ventricosum (DDNN) ×Triticum turgidum (AABB) hybrids and a single Triticum ventricosun×Triticum timopheevii (AABB) hybrid with colchicine. It would appear that the N-genome amphiploids can be utilized to transfer tolerance to aluminum to cultivated Triticum species and to study the genetics of tolerance in the N genome.     

Characterization of wheat-Thinopyrum partial amphiploids for resistance to barley yellow dwarf virus

Resistance to viruses such as wheat streak mosaic virus (WSMV) and barley yellow dwarf virus(BYDV) is lacking in the primary gene pool of wheat, and therefore resistance is being introgressed from wild relatives such as Thinopyrum species. Resistance to BYDV was found in partial amphiploids (2n = 8x = 56, consisting of 42 wheat and14 alien chromosomes) obtained in hybrids between wheat and both Th. intermedium and Th.ponticum. GISH analysis revealed that the alien genomes of all but one resistant partial amphiploid were heterogeneous consisting of different ratios of St, J^s and J genome chromosomes obtained from theThinopyrum parent. Translocated chromosomes consisting of Robertsonian, interstitial and terminal translocations between the different genomes were also detected. The tissue blot immunoassay showed that partial amphiploids having resistance could be inoculated with the virus but both virus multiplication and spread were completely blocked. This revised version was published online in August 2006 with corrections to the Cover Date.     

Relative Efficiency of Anther Culture and Chromosome Elimination Techniques for Haploid Induction in Triticale × Wheat and Triticale × Triticale Hybrids

Summary The study was undertaken to evaluate the relative efficiency of anther culture and chromosome elimination (by crosses with maize) techniques of haploid induction in intergenotypic triticale and triticale × wheat hybrids. For this, 15 triticale × wheat and 8 triticale × triticale F₁ hybrids were subjected to anther culture and were also simultaneously crossed with the `Madgran Local' genotype of maize (Zea mays L.) to induce haploids through the chromosome elimination technique. The haploid embryo formation frequency through the chromosome elimination technique was significantly higher in both, triticale × wheat (20.4%) and triticale × triticale (17.0%) F₁ genotypes, as compared to the calli induction frequencies through anther culture (1.6 and 1.4%, respectively). Further, four triticale × wheat and three triticale × triticale F₁ genotypes failed to respond to anther culture, whereas, all the F₁ genotypes formed sufficient number of haploid embryos through the chromosome elimination technique with no recovery of albino plantlets. The haploid plantlet regeneration frequencies were also significantly higher through the latter technique in both triticale × wheat (42.7%) and triticale × triticale (49.4%) F₁s as compared to anther culture (8.2 and 4.0%, respectively), where the efficiency was drastically reduced by several constraints like, high genotypic specificity, low regeneration frequency and albinism. The overall success rates of obtaining doubled haploids per 100 pollinated florets/anthers cultured were also significantly higher through the chromosome elimination technique (1.1% in triticale × wheat and 1.5% in triticale × triticale hybrids), proving it to be a highly efficient and economically more viable technique of haploid induction as compared to anther culture, where the success rates were only 0.2% and 0.1%, respectively.     

Cytology and Fertility of Hybrids and Amphiploids between Aegilops caudata L. ×Triticum turgidum (L.) Thell

Interspecific hybrids and amphiploids between Aegilops caudata L. (2n = 2x = 14, CC) and Triticum turgidum (L.) Thell. conv. durum Desf M. K. (2n = 4x = 28, AABB) were produced. Such hybrids can be used to introduce desirable traits such as disease resistance into cultivated durum wheats. One of the durum parents was a ph I mutation of the cv. ‘Cappelli’ used for testing the possibility of direct introduction of alien variation into cultivated species. The amphiploids were obtained both through colchicine chromosome doubling and as natural non-reductional mciosis products. In both hybrids and amphiploids, meiotic pairing and fertility were studied. Hybrids showed varying degrees of pairing and, in addition to the one involving the ph 1 mutant, one high pairing hybrid was found (Ae. caudata× cv. ‘Capinera’). Cytological examination of microsporogenesis in amphiploids revealed a high frequency of bivalent formation. Fertility proved to be a very variable character since some of the amphiploids were almost completely sterile. The use of amphiploids in breeding programmes is discussed in relation to meiotic and fertility data.     

The Attempted Transfer of Male Sterility from the Cornerstone Mutant of Tetraploid Wheat to Diploid Wheat

An attempt was made to transfer male-sterility, mslc, from the Cornerstone mutant of tetraploid wheat to diploid wheat. The N-banding technique revealed that chromosome 4 A of tetraploid wheat does not pair with chromosome 4 of diploid wheat in triploid F₁ hybrids: consequtntly the transfer of male-sterility gene(s) from tetraploid wheat to diploid was not successful. The culchiane induced ampliploids possessing AAAABB in mslc background were fully fertile indicating the complete compensation of mslc by the newly introduced A genome of T. monocaccum. The fertility compensating gene(S) presumably located on chromosome 4 of diploid wheat may be used to produce hybrid wheat by the XYZ system.     

Cytological analyses of hybrids and derivatives of hybrids between durum wheat and Thinopyrum bessarabicum,using multicolour fluorescent GISH*

The wheat progenitors and other wild relatives continue to be important sources of genes for agronomically desirable traits, which can be transferred into durum wheat (Triticum turgidum; 2n = 4x = 28; AABB genomes) cultivars via hybridization. Chromosome pairing in durum × alien species hybrids provides an understanding of genomic relationships, which is useful in planning alien gene introgression strategies. Two durum cultivars, ‘Lloyd’ and ‘Langdon’, were crossed with diploid wheatgrass, Thinopyrum bessarabicum (2n = 2x = 14; JJ), to synthesize F₁ hybrids (2n = 3x = 21; ABJ) with Ph1. ‘Langdon’ disomic substitution 5D(5B) was used as a female parent to produce F₁ hybrids without Ph1, which resulted in elevation of pairing between durum and grass chromosomes – an important feature from the breeding standpoint. The F₁ hybrids were backcrossed to respective parental cultivars and BC₁ progenies were raised. ‘Langdon’ 5D(5B) substitution × Th. bessarabicum F₁ hybrids were crossed with normal ‘Langdon’ to obtain BC₁ progeny. Chromosome pairing relationships were studied in F₁ hybrids and BC₁ progenies using both conventional staining and fluorescent genomic in situ hybridization (fl‐GISH) techniques. Multicolour fl‐GISH was standardized for characterizing the nature and specificity of chromosome pairing: A–B, A–J and B–J pairing. The A–J and B–J pairing will facilitate gene introgression in durum wheat. Multicolour fl‐GISH will help in characterizing alien chromosome segments captured in the durum complement and in their location in the A and/or B genome, thereby accelerating chromosome engineering research.     

Anther culture response of F1 durum × bread wheat hybrids after colchicine application

The effect of colchicine on the androgenic response of durum × bread wheat F₁ hybrids was studied. For this, three Greek durum wheat cultivars, exhibiting good yield and pasta quality, were crossed to two bread wheat cultivars with good response to anther culture. Spikes of the resulting hybrids, containing microspores in the mid to late uninucleate stage, were selected and cultured on two different media (W₁₄ and solid potato‐2) with and without colchicine. A negative effect on anther culture response was observed when colchicine was added to the W₁₄ induction medium. A similar effect was observed in two of the hybrids when they were cultured on to potato‐2 induction medium. In both induction media, green plant production was influenced negatively by colchicine treatment. The same was observed in albino plant production. The results of the present study support the view that anther culture response is strongly genotype dependent. Finally, potato‐2 induction medium was the most suitable one for the material studied.     

The association of genes for purple coleoptile with chromosomes of the wheat variety Mironovskaya 808

Summary The association of genes for purple pigment in the coleoptile with the chromosomes of the winter wheat variety Mironovskaya 808 was investigated using monosomic F₂ analysis. The segregation ratio for F₂ hybrids of Chinese Spring monosomics x Mironovskya 808 seems to indicate that the purple colour of the coleoptile is determined by two dominant genes, Rc₃ and Rc₄, which are located on the chromosomes 7D and 6B respectively, and which reinforce each other. Apart from these two genes, suppressors found on the chromosomes 2A, 2B, 2D, 4B and 6A also play a role in the intensity of the purple colour.With the aid of a Chinese Spring telocentric chromosome marker it was observed that the Rc₃ gene is located on the chromosome arm 7DS, at a distance of 16±4.23 crossover units from the centromere.     

Genotypic control of chromosome pairing in amphiploids involving the cultivated oat Avena sativa L.

Summary A genotype of the diploid species Avena longiglumis (Cw 57) has been shown to modify the genetic control of diploid-like chromosome pairing in the cultivated oat, A. sativa (2n=6x=42) leading to increased homoeologous chromosome pairing in 4x hybrids between the two species (Rajhathy & Thomas, 1974). The Cw 57 genotype has a similar effect in increasing homoeologous chromosome pairing in amphiploids combining diploid and hexaploid genomes including associations between alien chromosomes and their corresponding pairs in hexaploid species. The effect of the Cw 57 genotype is probably in altering the specificity of chromosome pairing in the early stages of meiosis. The use of the Cw 57 genotype to induce homoeologous chromosome pairing as a technique for the transfer of desirable alien variation into the cultivated oat is discussed.     

Introgression of Genetic Information from Triticum monococcum L, into Hexaploid Triticale by Hybridization with a T. monococcum×S. cereale Amphiploid

Four hexaploid triticale lines were crossed as females with a T. monococcum×S. cereale amphiploid (A^mA^mRR), with the aim of introducing the genetic material of diploid wheat. F₁-plants (A^mABRR)were back-crossed with a parental form of 6×-triticale as male and progenies were subjected to four different types of pollination with the aim of finding the optimal one in respect to gradual stabilization of introgressive hexaploid karyotypes. Beginning with BC₁-plants, a strong tendency to decrease the somatic chromosome number was observed. In subsequent generations this was accompanied by the decrease of seed germination and plant fertility. Both of these characters showed statistically significant dependence on somatic chromosome number variation which was analyzed in BC₁/F₂ and BC₂ populations. Based on spike fertility, an effective selection pressure was applied to restitute the hexaploid chromosome number. In the BC₁/F₄ generation, the first morphologically uniform secondary hexaploid lines were selected. 11.4% of the lines showed a non-waxy spike — a morphological marker transmitted from T. monococcum.