Assessment of genetic and phenotypic variation among intraspecific somatic hybrids of potato, Solanum tuberosum L.

Intraspecific somatic hybrids have been produced by protoplast fusion in eight combinations involving 10 dihaploids (2n= 2x= 24) in an attempt to provide new material for potato breeding. Cytological analysis revealed extensive variation in chromosome number, such as aneuploid, aneusomatic and mixoploid hybrids. Most of the hybrids represented the expected chromosome number of 48; however, the frequency of aneuploids reached 50% in some combinations. Some hybrids carried structurally rearranged chromosomes and exhibited a high frequency of aberrant anaphases. Isozyme and random amplified polymorphic DNA (RAPD) patterns of the hybrids from the same fusion combination were uniform. In the field, somatic hybrids showed wide phenotypic variation in 20 morphological characters. There was a significant correlation between certain leaf characters and the ploidy level, which may be used to distinguish the tetraploid hybrids from hexaploids and octoploids. Tuber yield and flowering intensity were highest in tetraploid hybrids (2n= 4x= 48). Eighteen of the 73 hybrids reached higher yields than the standard variety ‘Adretta’. Floral development and fertility were restored in hybrids derived from fusions between non-flowering or sterile dihaploids.     

Crosses between Hordeum vulgare L. and H. bulbosum L. I. Production,morphology and meiosis of hybrids,haploids and dihaploids

Summary From crosses between diploid and autotetraploid cytotypes of Hordeum vulgare L. (cultivated barley) and H. bulbosum L. (bulbous barley grass) diploid, triploid and tetraploid interspecific hybrids were produced. Both directly and after vegetative segregation crosses in either direction also gave rise to haploids and dihaploids resembling H. vulgare. The use of embryo culture was necessary. Plant morphology of the hybrids was much like that of H. bulbosum, although the hybrid plants were less vigorous. Meiosis in the hybrids was more or less disturbed, and this seemed to be the main cause of the high level of sterility.     

Fingerprinting of alfalfa meiotic mutants using RAPD markers

Summary A calendar of female sporogenesis and gametogenesis was made for both apomictic tetraploid (2n=4x=36) Brachiaria brizantha and Brachiaria decumbens and their apomictic F₁ hybrids with sexual tetraploid (2n=4x=36) Brachiaria ruziziensis. Microgametogenesis was used as a reference. Apospory was facultative in both species and hybrids. Environmental conditions had variable effects on the level of apomixis according to each genotype. Ratios of segregation into sexuals and apomicts in the interspecific hybrids suggest an oligogenic determinism with dominant apomixis in the genus Brachiaria. Highly apomictic and partially male fertile hybrids were identified and will be used in an improvement program to transfer genes for apomixis into the sexual species B. ruziziensis.     

Breeding for quantitative resistance to potato cyst nematode (Globodera pallida) in tetraploid potatoes using dihaploids and unreduced gametes

Summary Dihaploids were produced from tetraploids resistant to potato cyst nematode (Globodera pallida (Stone)). High levels of resistance were found in the dihaploids and three were used to produce tetraploid progenies by crossing them with susceptible tetraploid cultivars. One dihaploid, PDH505, produced more highly resistant offspring than the other two, PDHs 417 and 418. The latter gave progenies whose levels of resistance were similar to those obtained from susceptible dihaploids crossed with resistant tetraploids.The differences between the progenies of the resistant dihaploids were probably due to different modes of unreduced gamete formation (PDH505 producing gametes by first division restitution (FDR) and PDHs 417 and 418 by second division restitution (SDR)) although cytological studies would be necessary to confirm this. The methods by which dihaploids could be utilised in a tetraploid potato breeding programme are discussed in relation to the mode of unreduced gamete formation.     

The Diploidised Meiosis of Tetraploid Beta macrocarpa and its Possible Application in Breeding Sugar Beet

Accessions of Beta macrocarpa Guss., collected on the Canary Islands, were tetraploid (2n = 36). Three of these accessions were studied in detail. The plants were rather uniform in their morphological appearance, both within and between accessions, and were annual and fully self-compatible. Meiosis was completely diploidised, suggesting an alloploid nature of the tetraploid species. Crosses with diploid B. vulgaris yielded triploid hybrids which were sterile or nearly so; a few descendants of such hybrids were highly aneuploid. Crosses between tetraploid B. macrocarpa and autotetraploid cytotypes of B. vulgaris showed variable results, only part of these crosses yielded tetraploid hybrids. The tetraploid hybrids exhibited somewhat higher fertility than the triploids. An F₂ generation showed partial hybrid dwarfness, partial fertility and segregation for carliness and coloration of the hypocotyl. All hybrids had multivalents at meiosis (averages: 2.5—5.4 multivalents per pollen mother cell), indicating suppression of the diploidised meiosis. The possibilities of application of the diploidised meiosis in breeding sugar beet are discussed.     

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.     

Fertility of Brachiaria ruziziensis in interspecific crosses with Brachiaria decumbens and Brachiaria brizantha: meiotic behaviour,pollen viability and seed set

Summary Colchicine induced tetraploid (2n=4x=36) Brachiaria ruziziensis were used as female parent in crosses with apomictic tetraploid species (2n=4x=36) Brachiaria decumbens and Brachiaria brizantha. Tetraploid B. ruziziensis pollinated with B. decumbens set significantly more seed than selfed or crossed with B. brizantha. The crossability between B. ruziziensis and B. decumbens is also better than between B. ruziziensis and B. brizantha. In addition, hybrid seedlings obtained in crosses involving B. brizantha are more frequently lethal. All the viable F₁ hybrids are tetraploid with 36 chromosomes. Meiotic chromosome behaviour suggest that the three species belong to the same genomic group and therefore the same agamic complex. Chromosome associations at metaphase I do not allow to identify fertile and sterile hybrids. The interspecific hybrids averaged a lower fertility than their female parent, but some hybrids were more fertile than their apomictic male parent.     

AFLP analysis of Solanum phureja DNA introgressed into potato dihaploids

Introgression of Solanum phureja DNA into S. tuberosum dihaploids was analysed by the use of amplified fragment length polymorphism (AFLP) markers. Five dihaploids, derived from crosses between S. tuberosum cv. Pentland Crown and two different S. phureja pollinators (IVP48 and EC90) were investigated by use of 17 AFLP primer pairs. Also 30 dihaploids, derived from pollination of five different S. tuberosum seed parents with S. phureja IVP101, were investigated for the presence of S. phureja‐specific markers. In total approximately 680 and 850 AFLP products were detected in the diploid S. phureja clones and in the tetraploid S. tuberosum genotypes, respectively. A total of 68 S. phureja IVP48‐specific markers were detected, while the total number of S. phureja IVP101‐specific markers was in the range of 72‐96, depending on the S. tuberosum seed parent. Introgression of DNA in the S. tuberosum cv. Pentland Crown dihaploids, after pollination with S. phureja IVP48 and S. phureja EC90, was demonstrated by the detection of 14 of 68 IPV48‐specific markers in the dihaploids. However, no DNA introgression was found in any of the 30 S. tuberosum dihaploids derived from S. phureja IVP101. Hence, S. phureja IVP101 is regarded as an excellent pollinator in the production of S. tuberosum dihaploids in potato breeding programmes because of the high yield of dihaploids per 100 berries, and because no introgression of DNA into the S. tuberosum dihaploids was evidenced.     

Meiosis in different F1-hybrids of Solanum acaule Bitt. x S. bulbocastanum Dun. and its bearing on genome relationship,fertility and breeding behaviour

Summary Meiosis was studied in some triploid, tetraploid and hexaploid F₁-hybrids from Solanum acaule x S. bulbocastanum and in a triploid F₁-hybrid from S. acaule x S. tuberosum-haploid.The only anomaly found was stickiness at metaphase I, the degree of which appeared to be related to the proportion of the S. bulbocastanum chromosomes in the hybrids. No stickiness occurred at pre- and post-metaphase stages.A clear allosyndetic pairing between chromosomes of the two S. acaule genomes was observed in all triploid and tetraploid hybrids. These genomes apparently are closely related and little differentiated. The triploids S. acaule x S. bulbocastanum had 0–1 trivalent per cell, whereas 2–5 trivalents per cell were observed in the triploid S. acaule x S. tuberosum-haploid. Therefore S. acaule is more closely related to S. tuberosum L. than to S. bulbocastanum. A small amount of pairing between S. acaule and S. bulbocastanum is apparent from the occurrence of multivalents in all hybrids.Hexaploid F₁'s (2n=72) showed a nearly complete homologous pairing of chromosomes (35.2 bivalents per cell) and generally a normal separation of chromosomes at anaphase: 36-36. This offers an explanation for their high fertility. Triploid F₁'s from S. acaule x S. bulbocastanum have a high frequency of univalents leading to irregular separation of chromosomes at anaphase and consequently to unbalanced gametes and extreme sterility. In the tetraploid F₁'s (2n=48) nearly complete bivalent pairing was observed, 50% expectedly being homologous and 50% homoeologous pairing. Separation of chromosomes at anaphase was generally normal 24-24. In spite of this normal behaviour and allowing for tight stickiness at metaphase the tetraploids are very sterile. A satisfactory explanation cannot yet be given.Selfing and intercrossing hexaploid F₁'s gives normal berry set and many seeds per berry. However crosses with S. tuberosum and even those with the fertile hexaploid hybrid from 8x-S. acaule x 4x-S. tuberosum are little successful: berry set is far below normal and the berries are either parthenocarpic or contain only one or two seeds. These rather unexpected results warrant further investigation. Large-scale selfings and intercrosses of triploid and tetraploid hybrids have not been successful as yet. Among the female gametes of tetraploid hybrids a few appeared to be functional in crosses with hexaploid hybrids and in those with S. bulbocastanum.     

Monohaploids (n=x=12) from autotetraploid Solanum tuberosum (2n=4x=48) through two successive cycles of female parthenogenesis

Summary Viable monohaploids with 12 chromosomes in their somatic cells were obtained from an autotetraploid cultivar of Solanum tuberosum (2n=4x=48) by inducing female parthenogenesis successively in the tetraploid cultivar and in dihaploids from that cultivar. Both dihaploids and monohaploids were induced using the diploid S. phureja clones IvP 35 and IvP 48 as pollinators. The average frequencies of dihaploids and monohaploids in 1973 were 2.0 and 0.14 per berry respectively.Non-homologous chromosome associations (bivalents, trivalents and even quadrivalents) were observed at metaphase I of meiosis in pollen mother cells of the two monohaploids studied.The occurrence of non-homologous associations of chromosomes during meiosis is discussed. In addition the potential significance of monohaploids for basic research andfor breeding of potatoes is considered.     

Introgression of Cajanus platycarpus genome into cultivated pigeonpea, C. cajan

Summary Cajanus platycarpus, an incompatible wild species from the tertiary gene pool of pigeonpea (C. cajan (L.) Millspaugh), has many desirable characteristics for the improvement of cultivated varieties. To necessitate such transfers, embryo rescue techniques were used to obtain F₁ hybrids. The F₁ hybrids were treated with colchicine to obtain tetraploid hybrids, that were selfed to obtain F₂, F₃ and F₄ progenies. All of the hybrids and subsequent progenies had an intermediate morphology between the two parents. Backcrossing of the tetraploid hybrids with cultivated pigeonpea was not possible given embryo abortion, with smaller aborted embryos than those obtained in the F₀ parental cross.As a route of introgression, diploid F₁ hybrids were backcrossed with cultivated pigeonpea and BC₁ progeny obtained by in vitro culture of aborting embryos. BC₂ plants were obtained by normal, mature seed germination. Although embryo rescue techniques had to be used to obtain F₁ and BC₁ plants, it was possible to produce BC₂ and subsequent generations through direct mature seed. Every backcross to cultivated pigeonpea increased pollen fertility and the formation of mature seeds.Special project assistant till December, 2003.     

The transfer of triazine resistance from Brassica napus L. to B. oleracea L. II. Morphology,fertility and cytology of the F1 hybrid

Summary F₁ hybrids of triazine resistant Brassica napus and triazine susceptible B. oleracea were morphologically intermediate to the parent species. Of 49 hybrids examined, 44 had 28 chromosomes, two had 37, one had 38 and two had 56. The 38-chromosome plant was thought to be a matromorph, the others, A₁C₁C (28), A₁C₁CC (37) or A₁A₁C₁C₁CC (56) type hybrids. Pollen stainability averaged 9.0% in the sesquidiploid, 32.0% in the tetraploids and 89.5% in the hexaploids. All the interspecific hybrids were resistant to 1.0×10^-4 mol L^-1 atrazine. The sesquidiploid hybrids produced gametes with chromosome numbers ranging from 9 to 17 and the tetraploid hybrid gametes had chromosome numbers from 15 to 22. Most hybrids produced self-seed. The partial fertility of these hybrids may permit their backcrossing to one or both parents.     

Intra‐ and inter‐specific crosses of Solanum materials after mitotic polyploidization in vitro

Mitotic polyploidization in vitro was used in selected wild Solanum species and Solanum tuberosum dihaploids. The efficiency of polyploidization by colchicine was compared with that of oryzalin. Oryzalin was more effective than colchicine (P = 0.1). The rate of non‐affected to mixoploid to tetraploid regenerants was 22 : 2.5 : 1 (colchicine) and 14 : 2 : 1 (oryzalin). Optimal concentrations and durations were 3.5 mm /24 h for colchicine and 25 or 30 μm for 24 or 48 h for oryzalin (variations in concentration and duration are necessary owing to possible diversity of responses in selected genotypes). Tetraploids were obtained from S. berthaultii, S. bulbocastanum, S. pinnatisectum, S. verrucosum and eleven S. tuberosum dihaploids. The yield of tetraploids derived from tbr dihaploids was lower than that from the wild species (P = 0.01). Tetraploid regenerants were tested in intra‐ and inter‐specific crosses. Three of 43 intra‐specific combinations (298 pollinated flowers) were successful and yielded 440 seeds. Inter‐specific crosses (138 combinations, 1672 pollinated flowers) yielded 48 seedless berries.     

Tolerance to low temperatures and tuber soft rot in hybrids between Solanum commersonii and Solanum tuberosum obtained through manipulation of ploidy and endosperm balance number (EBN)

The objectives of this study were to evaluate the tolerance to low temperatures and tuber soft rot in hybrids between Solanum commersonii and Solanum tuberosum. The experimental materials consisted of F₁ triploid, BC₁ pentaploid‐near pentaploid and BC₂ tetraploid–near tetraploid hybrids. The F₁ triploids had a freezing tolerance and acclimatization capacity closest to S. commersonii. This indicated that the endosperm barriers which prevent the introgression of 1EBN S. commersonii into 4EBN S. tuberosum had been overcome. Indeed, the triploids produced 2n eggs, thus giving a compatible maternal to paternal EBN ratio in the hybrid endosperm generated by the 3x(2EBN) × 4x(3EBN) crosses. The tolerance to low temperatures of BC₁ and BC₂ hybrids was lower than that of the F₁. However, a number of genotypes were identified which were able to withstand temperatures down to ‐5°C. Some BC₂ hybrids were also tested for their tolerance to tuber soft rot, and some resistant hybrids were detected. A number of them combined the capacity for cold acclimatization with tolerance to tuber soft rot. These hybrids have an EBN of 4; they are fertile and have been used in backcrosses with 4EBN S. tuberosum.     

An assessment of the fertiliser response and other characters of three dihaploid potatoes and the characteristics of their tetraploid progenies

Summary Three dihaploids, two tetraploid cultivars and an unnamed tetraploid potato were subjected to three fertiliser treatments in a glasshouse. One dihaploid (PDH135), as well as being high yielding, responded to fertiliser dosage in a similar way to the cultivars by increasing tuber yield with each increase in fertiliser. The other two dihaploids increased shoot growth rather than tuber yield.An evaluation of tetraploid offspring obtained from dihaploid × tetraploid crosses, showed that PDH135 had the highest-yielding progenies in field trials. Other characters of the dihaploids were efficiently transferred into their tetraploid offspring. The use of dihaploids and unreduced gametes in breeding cultivars with useful quantitative characters is discussed.     

Breeding for low seed caffeine content of coffee (Coffea L.) by interspecific hybridization

Summary Seeds from open-pollinated flowers collected from hybrids of several Coffea species were analysed for caffeine content. The caffeine content was not always intermediary to that of the parents; higher and lower values were found. Diploid F₁ hybrids between accessions of C. eugenioides and C. salvatrix showed the lowest seed caffeine content. Seeds of the tetraploid hybrids C. arabica × C. salvatrix or C. arabica × C. eugenioides hybrids presented low caffeine content. The possibility of breeding coffee to reduce the caffeine content in the seeds by interspecific hybridization of C. arabica with other Coffea species is discussed.     

Genetics of self-compatibility in dihaploids of Solanum tuberosum L. I. Breeding behaviour of two self-compatible dihaploids

Summary Two highly fertile and self-compatible dihaploids (2ns=2x24) from Solanum tuberosum L. (2n 4x 48) were investigated to elucidate the genetic basis of their self-compatibility. To this end the two dihaploids were selfed and reciprocally intercrossed and the resulting I₁ and F₁ plants tested for self-compatibility. Reciprocal backcrosses of I₁-plants and F₁-plants were made. Complete diallels both within self-compatible and within self-incompatible F₁-plants were carried out as well as reciprocal matings between self-compatible and self-incompatible F₁-plants. From the wealth of data it could be concluded, that the dihaploids have two intact S-alleles, one being common to both. Six hypotheses were tested for explaining self-compatibility in these particular dihaploids. All but one had to be discarded. It is concluded that the self-compatibility most likely is brought about by the presence of an S-bearing translocation, which is not linked to the S-locus. The ratio sc :si in the F₁'s point either to certative disadvantage of translocation-bearing pollen or to lethality of translocation homozygotes. The importance of this self-compatibility mechanism for genetic and breeding research in potato is discussed.     

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.     

Interspecific hybridization in sexual diploid Allium senescens var. minor× apomictic tetraploid A. nutans and sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens

Fourteen interspecific hybrids in sexual diploid Allium senescens var. minor× apomictic tetraploid Allium nutans L. crosses, and eight interspecific hybrids in sexual diploid A. senescens var. minor× apomictic hexaploid A. senescens L. crosses were produced. The number of chromosomes was 2n= 24 in interspecific hybrids of diploid × tetraploid, and 2n= 32 in diploid × hexaploid crosses. Triploid and tetraploid interspecific hybrids showed intermediate parental morphological characteristics. Tetraploid interspecific hybrids of A. senescens var. minor×A. senescens crosses formed two groups based on leaf colour and leaf width. Seeds were formed in 11 out of 14 triploid interspecific hybrids under natural conditions. In cytological observations of parthenogenesis, three out of 12 triploid interspecific hybrids and five out of eight interspecific tetraploid hybrids were observed. Parthenogenesis ranged from 26.0% to 86.0% in five tetraploid interspecific hybrids. Non-parthenogenesis to parthenogenesis segregated in a 3:5 ratio in A. senescens var. minor×A. senescens crosses.