Interfering with regular meiotic behaviour in Avena sativa as a method of incorporating the gene for mildew resistance from A. barbata

Summary The regular meiotic behaviour of the cultivated oat Avena sativa (2n=6x=42) is genetically controlled. The factors which control the diploid-like meiotic behaviour also restrict the amount of pairing that occurs between alien chromosomes and their homoeologues in A.sativa, and hence increases the difficulties of introducing desirable variation from wild species into the cultivated oat. A genotype of the diploid species A.longiglumis which interferes with the regular meiotic behaviour of A. sativa can be used to induce pairing between alien chromosomes and their corresponding chromosomes in A. sativa. Using this procedure the dominant gene conferring mildew resistance has been transferred from the tetraploid weed species A. barbata into the cultivated oat.     

A synthetic hexaploid (2n = 42) oat from the cross of Avena strigosa (2n = 14) and domesticated A. magna (2n = 28)

A synthetic hexaploid oat was produced by chromosome doubling of a sterile triploid hybrid between cultivated Avena strigosa (2n = 14) cv. Saia and a domesticated form of A. magna (2n = 28). The synthetic hexaploid was intermediate between its parents in panicle shape and lemma color, similar to the tetraploid parent in spikelet structure, and to the diploid parent in having a single, albeit partially shriveled seed per spikelet, and low protein content. By the third generation, plants with yellowish lemmas, mostly two seeds per spikelet and better filled grains had been selected. Rust resistance of the diploid parent was retained in the synthetic hexaploid, but not tolerance to barley yellow dwarf virus disease (BYDV). Chromosome associations at meiosis in the triploid hybrid was low, with over 60% of them being univalents. Bivalent association was the rule in the synthetic hexaploid with an occasional one or two quadrivalents. Regular meiosis with 21 bivalents was observed in further generations. The preferential pairing of homologous chromosomes in the synthetic hexaploid was probably contributed by the A. strigosa genome which exhibits this tendency in artificial allopolyploid situations. Selection of yellow lemma color and two seeds per spikelet suggests that the genes controlling these traits are located on the chromosomes involved in quadrivalents in the synthetic hexaploid. The potential and limitations of utilizing the synthetic hexaploid in oat research and breeding are briefly discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Discovery of 2n gametes in tetraploid oat Avena vaviloviana

Summary Sexual polyploidization via the action of 2n gametes (gametes with the sporophytic chromosome number) has been identified as the most important evolutionary mode of polyploidization among plant genera. This study was conducted to determine whether 2n gametes are present in the tetraploid level of the genus Avena (2n=4×=28) Twenty tetraploid Avena lines, representing four species and one interspecific hybrid, were screened for pollen grain size in order to differentiate between n and 2n pollen. Avena vaviloviana (Malz.) Mordv. line PI 412767 was observed to contain large pollen grains at a 1.0% frequency. Cytogenetic analyses of pollen mother cells of PI 412767 revealed cells with double the normal chromosome number (i.e., 56 chromosomes at metaphase I and anaphase I). The mode of chromosome doubling was found to be failure of cell wall formation during the last mitotic division that preceded meiosis. The resulting binucleate cells underwent normal meiotic divisions and formed pollen grains with 28 chromosomes. Based on the formation and function of 2n gametes, three models involving diploid and tetraploid oat lines are proposed to describe possible evolutionary pathways for hexaploid oats. If stable synthetic hexaploid oat lines could be developed by utilizing 2n gametes from diploid and tetraploid oat species through bilateral sexual polyploidization, the resulting hexaploids could be used in breeding programs for transferring genes from diploids and tetraploids to cultivated hexaploids.     

Introgression of crown rust resistance from diploid oat Avena strigosa into hexaploid cultivated oat A. sativa by two methods: direct crosses and through an initial 2x·4x synthetic hexaploid

New sources of resistance to crown rust, Puccinia coronata f. sp. avenae (Eriks.), the major fungal disease of cultivated oat, Avena sativa L. (2n = 6x = 42), are constantly needed due to frequent, rapid shifts in the virulence pattern of the pathogen. Crown rust resistance identified in the diploid oat A. strigosa (Schreb.) (2n = 2x = 14) accession CI6954SP was transferred into cultivated oat using two methods: direct cross of the diploid to a hexaploid cultivar facilitated by embryo rescue, and initial cross of the diploid to a wild tetraploid oat to make a synthetic hexaploid for subsequent crossing to a hexaploid cultivar. Two tetraploids, a crown rust resistant A. murphyi (Ladiz.) accession P12 and a susceptible A. insularis (Ladiz.) accession INS-1, were used in the 2x·4x crosses. Resistant backcross-derived lines were recovered by both methods. Although the 2x·4x synthetic method did not require the laborious discovery and rescue of an infrequent initial hybrid embryo of the direct cross, the direct cross method provided more rapid backcross recovery of plants with high fertility, full transmission of resistance, and desired plant and seed phenotypes. A suppressor effect, present initially but segregating in backcrosses, appeared to come from the CI6954SP donor and is the same as, or analogous to, suppression by an oat line with the crown rust resistance gene Pc38. No resistance from A. murphyi P12 was detected in advanced generations when it was introduced either as a component of a synthetic hexaploid or in direct crosses to A. sativa, indicating suppression of its resistance in interploidy combinations. That the dominant resistance gene transferred from CI6954SP and a gene Pc94 introgressed earlier from a different A. strigosa accession may be the same or quite similar to one another is indicated by their in-common specificity to suppression of resistance expression, susceptibility to a newly recovered rust isolate, and close linkage to the molecular marker SCAR94-2. The introgressed resistance genes from the different sources, even if the same, may have different cultivar genomic introgression sites, which would allow tests of dosage effects on resistance expression.     

Transfer of high kernel weight and high protein from wild tetraploid wheat (Triticum turgidum dicoccoides) to bread wheat (T. aestivum) using homologous and homoeologous recombination

Summary Wheat pentaploids were produced by hybridizing a high kernel weight (1000 grain wt=56 g), high protein (25.4%) line of wild tetraploid wheat (Triticum turgidum dicoccoides) as male parent, with the three hexaploids (T. aestivum) — normal Chinese Spring and its two homoeologous pairing mutants, ph _1b and ph ₂. The pentaploids were crossed as female parents to the two commercial hexaploid cultivars Warigal and Barkaee and 42-chromosome stable plants selected from the F₁ of the pentaploid x hexaploid crosses.Mean protein content of certain F₃ lines from all six pentaploid x hexaploid crosses was significantly higher than Chinese Spring and the respective commercial hexaploid parent (p<0.005) indicating high protein had been transferred from the tetraploid to the hexaploid level.Kernel weight amongst certain F₃ lines of the three pentaploids x Barkaee was significantly (p<0.0005) higher than either Chinese Spring or Barkaee, indicating the transfer also of high kernel weight from the tetraploid to the hexaploid level. However kernel weight was not significantly increased over Warigal in any F₃ lines of its crosses with the three pentaploids.High levels of homoeologous chromosome pairing in the ph-mutant pentaploids, plus evidence for significant modification of the composition of high-molecular weight (HMW) glutenin subunits of grain protein in certain F₃ derivatives of the ph-mutant pentaploid x hexaploid, crosses indicates that the ph-mutant-derived lines may possess novel (intergenome) genetic recombination, at least for high protein, and possibly kernel weight.     

Gene transfer from non-tuberous to tuberous Solanum species: Evidence from meiosis in hybrids

Summary The distant hybrids between non-tuberous Solanum species and tuberous S. pinnatisectum display little or no pairing in F1 and predominantly bivalent formation (preferential pairing) after chromosome doubling. In such a situation the question about the potential and extent of gene transfer from the non-tuberous parent to the tuberous one is relevant to potato breeding. This question was investigated by studying meiosis in triploid and hexaploid hybrids from crosses between diploid TV⁵ x tetraploid (S. etuberosum x S. pinnatisectum). TV⁵ is similar to S. verrucosum with cytoplasm of S. tuberosum. The following evidence was found for the desirable transfer of S. etuberosum genes to the tuberous species.The triploid F1 hybrids did not display the configurations 12 II+12 I expected if no gene exchange would take place between S. etuberosum and the tuberous species; however, a considerable number of multivalents per cell was observed in all plants studied.In the hexaploid F1 hybrids, obtained from the triploids through somatic doubling in vitro, 36 bivalents could reasonably be expected. Although bivalents were predominant (an overall average of 24.2 per cell) quite a few chromosomes were associated as multivalents in all plants investigated.It is concluded that in the hybrids studied a considerable amount of pairing and chiasma formation occurs between chromosomes of non-tuberous and those of tuberous Solanum species. This pairing affinity is larger than that found in 2x and 4x hybrids from S. etuberosum x S. pinnatisectum. Some hypotheses are put forward to explain this increased pairing affinity.     

Alien chromosome transmission and somatic elimination in monosomic addition lines of Gossypium australe F. Muell in G. hirsutum L.

The efficiency of using monosomic alien addition lines (MAALs) to introgress agronomical traits of interest carried by wild diploid Gossypium species into the main cultivated cotton species G. hirsutum depends on the opportunities of confronting the alien chromosome with the recipient background genome at each generation and on the occurrence of translocations and homoeologous recombinations. The selfed-progeny of five MAALs of G. australe in G. hirsutum was screened with SSR markers to determine the transmission frequency of the alien chromosome and monitor its integrity. Three MAALs revealed a transmission frequency significantly lower than the expected ratio and one MAAL presented an exclusive transmission of the additional chromosome. In these four MAAL the alien chromosome was transmitted almost unaltered. With the fifth MAAL the alien chromosome was normally transmitted but was altered in half of the plants carrying it. In one MAAL, normally carrying brown fiber, the emergence of some plants carrying white and brown fiber revealed the somatic elimination of the additional chromosome. The loss of this chromosome seems to be triggered by its deletion.     

Breeding for hard red winter wheat cultivars adapted to conventional-till and no-till systems in northern latitudes

Summary Monosomic alien addition lines combining individual F. drymeja chromosomes and the L. multiflorum complement were isolated from the cross between the triploid hybrid L. multiflorum (4x) × F. drymeja (2x) and diploid L. multiflorum (2x). Chromosome pairing in the addition lines was studied at metaphase 1 of meiosis and the relationship between single F. drymeja chromosomes and the corresponding homologous pair in L. multiflorum is discussed. Trivalent frequency in the addition lines was higher than expected from observations of chromosome pairing in the triploid hybrid and there were differences between lines in the number of trivalent associations formed. There is some evidence to suggest that trivalent frequency is not entirely dependent on chromosome length and that transmission of the alien chromosome in the female is dependent on the size of the added chromosome. Morphological studies were made to assess the phenotypic effects of the addition of single F. drymeja chromosomes to the L. multiflorum complement. Two plants (2n=14) with recombination between a L. multiflorum and a F. drymeja chromosome were identified.     

Evaluation of the use of species hybrids and synthetic amphiploids in the improvement of the oat crop

Summary Cytogenetic studies on species hybrids and synthetic amphiploids in the Avenae are discussed in relation to their use in transferring variation from wild species of different ploidy into cultivated hexaploid oats. Procedures for the introduction of alien variation into the cultivated oat are proposed, based on our present knowledge of the genetic relationships between species in Avena.     

Hybrids of <Emphasis Type="Italic">Avena sativa</Emphasis> with two diploid wild oats (CIav6956) and (CIav7233) resistant to crown rust

Two diploid accessions of wild oat, CIav6956 and CIav7233, were identified as carrying seedling resistance to oat crown rust (caused by Puccinia coronata f. sp. avenae; Pca). Two vigorous interploidy F₁ hybrids were generated from crosses involving the hexaploid oat cultivar Wintaroo and the diploid oat Avena strigosa Schreb. accession CIav6956. An additional interploidy F₁ hybrid, designated “F1-Aa1”, was produced from a cross of Wintaroo and the diploid oat accession CIav7233. All three hybrids were more vigorous and taller than the cultivated parent Wintaroo. The three F₁ hybrids contained full chromosome complements from both parents (2n = 4x = 28), but no seeds were obtained when the three F₁ hybrids were selfed. Meiotic analyses of the hybrids indicated that they exhibited a high degree of inter-genome and intra-genome pairing. Trivalent configurations were detected in 95–96% of meiotic cells and a minimum of three bivalents was present in all cells. An average chiasma frequency of 7.2–7.9 per cell was observed for the three F₁ hybrids. A fourth F₁ hybrid was subsequently generated from a cross between the diploid oat accession CIav7233 and Wintaroo. One octaploid (2n = 8x = 56) was generated from this hybrid and progeny were resistant to two Pca races. The chromosome number of the octaploid progeny varied between 51 and 54 chromosomes. Development of a chromosome addition line(s) with the crown rust resistance should be possible from these partial-octaploids.     

Ploidy Manipulation and Introgression of Resistance to Alternaria Helianthi from Wild Hexaploid Helianthus Species to Cultivated Sunflower (H. annuus L.) Aided by Anther Culture

The present investigation discusses the scope for transferring of resistance to leaf spot disease incited by Alternaria helianthi from two hexaploid wild species (H. tuberosus and H. resinosus) to diploid cultivated sunflower. Interspecific hybrids produced between sunflower and these two hexaploid species were partially fertile with tetraploid chromosome status. Backcrosses of these interspecific hybrids with cultivated sunflower resulted in the formation of sterile triploid plants. To overcome the problem of sterility and facilitate backcrosses with cultivated sunflower, anther culture of the tetraploid interspecific hybrids was carried out to bring down their chromosome number to diploid status. Anthers from both interspecific hybrids were cultured on basal Murashige and Skoog media supplemented with varying concentrations of organics and the growth regulators benzyladenine and naphthaleneacetic acid. Anthers of interspecific hybrids involving H. resinosus responded well and regenerated through an embryogenic route at a frequency of 98.7%. But in interspecific hybrids with H. tuberosus, anthers formed callus and subsequently regenerated shoots through an organogenic pathway. DNA ploidy analysis of anther culture plants of interspecific hybrids derived from H. tuberosus crosses was carried out to identify plants with desired diploid status. In vitro screening of parents, interspecific hybrids and anther culture plantlets against A. helianthi showed resistance in 68.5% of the anther culture plants of interspecific hybrids from H. tuberosus and in 24.3% of the plants derived from interspecific hybrids involving H. resinosus.     

An evaluation of intergeneric hybridization of Fragaria spp. x Potentilla spp. as a means of haploid production

Summary Unidirectional intergeneric crosses using octoploid and hexaploid Fragaria species as a female parent and diploid and tetraploid Potentilla species as a male parent were performed, producing in toto 1345 fruits with an estimated 46089 seeds. The germinating seeds obtained by using P. anserina mostly perished at the cotyledon stage, a few developing into matroclinous octoploid seedlings. The seeds obtained by using P. fruiticosa produced two types of seedlings: matroclinous octoploids and genuine intergeneric hybrids. It is considered that the matroclinous octoploids resulted from contamination during the emasculation process. A great majority of the hybrid seedlings were slow in growth and development and perished before reaching the flowering stage. No haploids have been recovered. Possible reasons for absence of haploid genotypes in the progenies of these intergeneric crosses are considered. Among the seedlings of F. x ananassa cv. Tioga x P. fruiticosa, a seedling of an unusual phenotype was noticed, which upon chromosome determination, proved to be 2n=9x=63. This genotype displayed complete male and female sterility. The implication of the sterility of this seedling for bridging of the two genera is discussed.This research was performed as part of a cooperative project on genetic improvement of cultivated strawberry between the Fruit Laboratory, USDA, Beltsville and NJAES. Paper of the Journal Series, D-12102-16-83. New Jersey Agricultural Experiment Station, Rutgers University, New Brunswick, NJ 08903.     

Chromosome numbers and microsporogenesis in Brachiaria brizantha (Gramineae)

The genus Brachiaria, native to the African tropical savannas, has achieved significance as a pasture grass in many tropical and subtropical countries, including Brazil. Many species and accessions are polyploid and apomictic, which complicates the improvement of breeding stocks through hybridization. In support of breeding programs, cytogenetic characterization, including chromosome counts and evaluation of the meiotic behavior in the accessions of the Brachiaria has been undertaken at the Embrapa Beef Cattle Center. In this study, 22 accessions of B. brizantha were analyzed of which one was found to be diploid (2n = 2x = 18), 18 were tetraploid (2n = 4x = 36) and three were hexaploid (2n = 6x = 54). The meiotic chromosome behavior was slightly irregular in the diploid and in some tetraploid accessions, and highly irregular in most tetra- and hexaploid accessions. Meiotic abnormalities were those common to polyploidy, i.e., multivalent chromosome association at diakinesis and irregular chromosome segregation leading to micronuclei formation in the tetrad stage. Low frequencies of multivalent chromosome associations among polyploids suggest that they may be segmental allopolyploids. This revised version was published online in August 2006 with corrections to the Cover Date.     

The production of multispecific octoploids from Fragaria species and the cultivated strawberry

Summary A method of incorporating genetic material from five species of Fragaria and the cultivated strawberry into fertile octoploids is described. A synthetic octoploid derived from a hexaploid x diploid hybrid was crossed to octoploid cultivar breeding lines until a BC₂ hybrid was produced. A second synthetic octoploid in which two diploid species and a tetraploid species were combined was crossed to a cultivated strawberry to produce a hybrid breeding clone. The two breeding clones were crossed and 222 seedlings were produced. The seedlings were generally vigorous and fruitful, some having commercial potential.     

Polyploidy and sexual polyploidization in the genus Vaccinium

Genus Vaccinium,consisting of blueberries, cranberries, lingonberries and many related wild species, includes diploid, tetraploid, and hexaploid species. Most evidence indicates that the tetraploid species are autotetraploids, with non-preferential bivalent chromosome pairing. Although homoploid interspecific crosses usually produce numerous fertile hybrids if the parents are from the same section of the genus, inter sectional crosses at the diploid level normally produce no seedlings, weak seedlings, or seedlings that are very low in fertility. There is a strong but not complete triploid block within Vaccinium. Even with insections, tetraploid × diploid (and the reciprocal) crosses normally give only a few tetraploid and a few triploid hybrids. Hexaploid × diploid crosses within sections are very hard to make, and the few hybrids that have been obtained are pentaploid. The frequency of 2n gametes varies,both among genotypes within species and among species. Vaccinium pollen is normally shed in tetrads, and the frequency of large pollen grains shed in dyads can be used to estimate 2n gamete frequency. Cultivated blueberries occur at both the tetraploid and the hexaploid levels, and there are important genetic resources in the diploids. Unreduced gamete production has permitted transfer of genetic material from the diploid to the tetraploid level and from the diploids and tetraploids to the hexaploid level via triploid hybrids. Intersectional crosses can occasionally produce tetraploid Vaccinium hybrids that appear to behave as amphidiploids and have medium to high fertility. CommercialVaccinium cultivars are normally propagated by cuttings. Intersectional hybridization, chromosome doubling, and asexual propagation could permit the production of novel hybrid combinations with value as ornamentals or in fruit production. This revised version was published online in July 2006 with corrections to the Cover Date.     

The production and behaviour of hexaploid hybrids between Hordeum vulgare and H. Bulbosum

Summary To produce hexaploid (or other polyploid) hybrids, diploid or tetraploid Hordeum vulgare was crossed with hexaploid or octoploid H. bulbosum, and perennial triploid hybrids between the two species were treated with colchicine. The crosses did not yield viable plants: seedset was low, the seed aborted and embryo culture was unsuccessful. The colchicine treatments geve rise to plants in which hexaploid chromosome numbers were observed. At the hexaploid level chromosomal instability occurred, resulting in chromosome elimination.The colchicine-treated triploid hybrids showed in the first years after the treatment better fertility after open flowering than untreated plants, but the level of fertility remained very low. The offspring consisted of haploid, diploid and approximately triploid plants like H. vulgare, tetraploid and approximately tetraploid plants like H. bulbosum, and plants with hybrid morphology and unstable chromosome number, which were highly sterile. Thus the crossing barrier between H. vulgare and H. bulbosum could not be broken down at higher ploidy level.     

Variation for crossability in a reciprocal,interspecific cross involvingHordeum vulgare andH. lechleri

Summary Several crossing series including the hexaploid (2n=6x=42), South American speciesHordeum lechleri and diploid (2n=2x=14) cultivated barley (H. vulgare) were performed. Barley functioned better as the paternal than as the maternal parent in all cases. Viable offspring were only obtained from one hybrid combination when barley was used as the maternal parent. There was an environmental influence on the success of crosses. A high seed set was counteracted by a lower germination frequency. The outcome is that different crossing series give similar results. As a result of chromosome elimination and in a few cases duplication of especially the barley chromosomes, the chromosome numbers of the offspring (239 plants) varied from 2n=21 (trihaploids ofH. lechleri) to 2n=30. Fifty-five % of the plants were euploid with the number expected for a hybrid (2n=28). The frequency of hyperploids, euploids, hypoploids (2n=22–27) and trihaploids varied by year, locality, type (winter/spring) of barley,H. lechleri population, and crossing direction.     

Somatic chromosome elimination and meiotic chromosome pairing in the triple hybrid 6x-(Solanum acaule × S. bulbocastanum) × 2×-S. phureja

Summary Two selected hexaploid F₁ clones from the cross Solanum acaule x S. bulbocastanum were intercrossed and the resulting hybrid plants pollinated with the diploid species S. phureja, in order to obtain tetraploid triple hybrids with the same ploidy level as S. tuberosum cultivars (2n=4x=48).Apart from three trihaploids a large population of triple hybrids was obtained, showing chromosome mosaicism in root tip cells (euploid + hypoploid chromosome numbers) and a uniform, mostly hypoploid chromosome number in the pollen mother cells. It is demonstrated that somatic chromosome elimination in the early stages of development is the most probable cause.From detailed pachytene observations as well as from the chromosome associations observed at metaphase I it is evident, that there is normal pairing between the four genomes in the triple hybrids. Although S. bulbocastanum is a quite distinct species with a rigid crossability barrier with S. phureja, the chromosomes of these two species appear to have a high degree of homology. Especially the formation of quadrivalents involving all twelve groups of four homeologous chromosomes, indicated that the four parental genomes (two from S. acaule, one from S. bulbocastanum and one from S. phureja) are not differentiated to the extent of affecting normal pairing and chiasma formation. These results support the view, that the transfer of valuable characters from S. bulbocastanum to S. tuberosum cultivars is feasible even when these characters are polygenically controlled.     

Interspecific hybridization in the genus Arachis through embryo culture

Summary The embryos of a cultivated tetraploid peanut (Arachis hypogaea), a wild diploid species (A. villosa), and their hybrid embryos, which generally abort in nature, were cultured in vitro and the plants have been successfully transferred to the soil. The hybrids showed triploid chromosome number (3x=30). The significance of wide hybridization in peanut-improvement programs is discussed.     

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.