Solanum phureja genes are expressed in the leaves and tubers of aneusomatic potato dihaploids

Summary The expression of leaf isozymes and tuber patatin in dihaploids derived from the Solanum tuberosum cv. Pentland Crown was investigated. Seven of the dihaploids were aneusomatic containing additional chromosomes from the S. phureja dihaploid inducer. Of these, four genotypes expressed leaf isozymes characteristic of the S. phureja dihaploid inducer, and the tubers of three aneusomatic dihaploids contained a S. phureja form of patatin. Aneusomatic dihaploids in which the proportion of cells containing additional S. phureja chromosomes was relatively small (i.e. 1–15%) did not express leaf isozyme markers or patatin bands characteristic of the dihaploid inducer or showed only faint expression of one or two markers. However, those with a high proportion of cells containing additional chromosomes (50–55%) had a range of strongly expressed leaf isozymes that were characteristic of the dihaploid inducer and also expressed the S. phureja tuber patatin.One dihaploid genotype was exclusively euploid (2n<24), yet is expressed a S. phureja leaf isozyme marker and S. phureja tuber patatin, suggesting recombination or chromosome substitution between the genome of the S. phureja dihaploid inducer and the cultivar Pentland Crown.     

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.     

The use of dihaploids in increasing the homozygosity of tetraploid potatoes

Summary Electrophoresis banding patterns of the tuber proteins of 12 dihaploids of the cultivar Pentland Crown showed that four had types of patatin unlike that of the parent. The patatin types of somatically chromosome-doubled clones derived from three of the dihaploids were identical to those of the dihaploid progenitors. Fourteen dihaploids produced from the chromosome-doubled derivative (PDH40X2) of one dihaploid, which had a variant patatin, had the patatin type of the parent dihaploid.The experiment showed that dihaploids and somatic chromosome doubling could be used to fix variation found in heterozygous tetraploid potatoes. Used together, haploidisation and chromosome doubling can generate highly homozygous tetraploids for use by plant breeders.     

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.     

Parthenogenetic monohaploids (2n=x=12) from Solanum tuberosum L. and S. verrucosum Schlechtd. and the production of homozygous potato diploids

Summary Dihaploid and dihaploid derived clones of Solanum tuberosum and diploid genotypes of S. verrucosum produced 85 viable monohaploids by female parthenogenesis. All were induced using diploid S. phureja clones, homozygous for embryo spot, as pollinator. Frequency of S. tuberosum monohaploids per 100 berries was rather constant in three successive years (14, 17 and 17 respectively). No male and female fertility was found in flowering monohaploids.Colchicine-induced chromosome doubling yielded homozygous s. tuberosum diploids with low pollen quality but good seed fertility.Two diploid self-incompatible species (S. multidissectum and S. berthaultii) produced no monohaploids. The presence of genes for female parthenogenesis in some dihaploids is discussed.     

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.     

Interspecific hybrids between three eggplant (Solanum melongena L.) cultivars and two wild species (Solanum torvum Sw. and Solanum sisymbriifolium Lam.)

Three Greek eggplant cultivars, ‘Langada’, ‘Tsakoniki’ and ‘Emi’ (2n= 24), were crossed with two wild species (Solanum torvum Sw., 2n= 24 and Solanum sisymbriifolium Lam., 2n= 24). Ovules isolated 15-27 days after pollination were cultured in a modified MS medium at 24°C and a 16h photoperiod. Fifty days later, the ovules were dissected and the interspecific embryos were cultured in the same medium. Interspecific hybrids were achieved only from crosses between the eggplant cultivars and S. torvum. The hybridity of the putative interspecific F¹ hybrid (Solanum melongena×S. torvum) was confirmed by using morphological and biochemical (isozyme isocitrate dehydrogenase A, phosphoglucomutase A, phosphoglucose isomerase B, 6-phosphogluconate dehydrogenase A, 6-phosphogluconate dehydrogenase B) markers. The F¹ plants (‘Langada’×S. torvum) were selfpollinated and backcrossed to both parents. Fruits, however, were produced only when the F¹ hybrid was backcrossed as female with the eggplant cultivar ‘Langada’.     

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.     

Induction of novel organelle DNA variation and transfer of resistance to frost and Verticillium wilt in Solanum tuberosum through somatic hybridization with 1EBN S. commersonii

Somatic hybridization can be used to induce genetic variability in plastidial and mitochondrial genomes, and transfer multiple uncloned genes across sexual barriers. Somatic hybrids were produced between a dihaploid clone of the common potato, S. tuberosum subsp. tuberosum, and the wild sexually incongruent diploid species S. commersonii. Fusion products were selected on the basis of callus growth and regeneration in vitro. Genome composition of putative somatic hybrids was determined by flow cytometric analysis of nuclear DNA content, RAPD analysis, and Southern analysis with probes specific to organellar DNA. All regenerated fusion products proved to be hybrids based on RAPD analysis. Seventy per cent of somatic hybrids were (near) tetraploids, 22% (near) hexaploids and 8%(near) octoploids. A high correlation was found between the nuclear DNA content determined by flow cytometry and the number of chloroplasts in stomata guard cell pairs. Somatic hybrids inherited the parental plastids in a random manner. On the contrary, they preferentially inherited the mitochondrial DNA fragments of S. tuberosum. The majority of them had a rearranged mitochondrial genome with fragments from both parents. Hybrids were highly vigorous and morphologically more similar to the cultivated than to the wild parent, produced tubers on long stolons under long photoperiod conditions, showed a high degree of flowering, but did not produce pollen. In addition, somatic hybrids were generally more resistant to frost and Verticillium wilt than the cultivated parent, indicating the introgression of relevant resistance genes from the wild species into the genetic background of S. tuberosum. This revised version was published online in July 2006 with corrections to the Cover Date.     

Analysis of dihaploid induction and production ability and seed parent x pollinator interaction in potato

Summary The production of dihaploids is the first step in a potato breeding program at the diploid level. Dihaploid induction ability, dihaploid production ability and seed parent x pollinator interaction were analyzed for 28 × 3, seed parent x pollinator combinations. This is the first report on significant interaction between seed parents and pollinators. Despite this interaction, IVP 101 had a significantly higher dihaploid induction ability than the widely used pollinators IVP 35 and IVP 48. Previous findings on significant differences in dihaploid production ability between seed parents were clearly confirmed.     

Identification of novel QTL for resistance to crown rot in the doubled haploid wheat population 'W21MMT70' × 'Mendos'

Crown rot (causal agent Fusarium pseudograminearum) is a fungal disease of major significance to wheat cultivation in Australia. A doubled haploid wheat population was produced from a cross between line ‘W21MMT70’, which displays partial seedling and adult plant (field) resistance to crown rot, and ‘Mendos’, which is moderately susceptible in seedling tests but partially resistant in field trials. Bulked segregant analysis (BSA) based on seedling trial data did not reveal markers for crown rot resistance. A framework map was produced consisting of 128 microsatellite markers, four phenotypic markers, and one sequence tagged site marker. To this map 331 previously screened AFLP markers were then added. Three quantitative trait loci (QTL) were identified with composite interval mapping across all of the three seedling trials conducted. These QTL are located on chromosomes 2B, 2D and 5D. The 2D and 5D QTL are inherited from the line ‘W21MMT70’, whereas the 2B QTL is inherited from ‘Mendos’. These loci are different from those associated with crown rot resistance in other wheat populations that have been examined, and may represent an opportunity for pyramiding QTL to provide more durable resistance to crown rot.     

Detection by simple sequence repeat markers of introgression from Coffea canephora in Coffea arabica cultivars

Microsatellites or simple sequence repeats (SSR) were used to assess polymorphism among 16 Coffea arabica and four Coffea canephora accessions, and to identify DNA introgression fragments from C. canephora in four C. arabica lines. Thirty‐one primer pairs allowed for the identification of 92 polymorphic alleles distributed over 37 loci. The C. arabica accessions derived from the genetic bases ‘Typica’ and ‘Bourbon’ were grouped separately according to their genetic origin. Two genotypes derived from a spontaneous hybrid (C arabica×C. canephora) were classified with the C. canephora accessions from Central Africa. Coffea canephora from West Africa were separated from the other accessions studied. Four alleles related to introgression (i.e. present in C. canephora and introgressed lines, and absent in C. arabica) were identified. The SSR markers were used successfully for characterization of a particular cultivar (‘Veranero’) from Costa Rica, which is known for its late maturity.     

Nuclear genome constitution and other characteristics of somatic hybrids between dihaploid Solanum acaule and tetraploid S. tuberosum

Eleven somatic hybrids (2n = 68 to 74) obtained between S. tuberosum ssp. tuberosum cv. Dejima (2n = 48) and ATDH-1 (2n = 24), an anther-culture-derived dihaploid of S. acaule (Yamada et al., 1997), were characterized by nuclear RFLP markers using 49 single-copy DNA probes distributed throughout the potato genome (2 to 6 probes per chromosome). One of the somatic hybrids, DA8-2, had 72 chromosomes and all the Dejima- and ATDH-1-specific markers (124 and 103 bands, respectively), suggesting the presence of a whole set of both parental chromosomes. The other somatic hybrids lost varying numbers of markers up to seventeen. The pattern of the loss of markers indicated the elimination of five chromosomes among four somatic hybrids. A nucleolar organizer region of chromosome 2 was often eliminated in the somatic hybrids. The somatic hybrids studied here had higher frequencies of multivalent formation than the S. tuberosum parent. They had reasonably good seed set when pollinated with S. tuberosum pollen. Hence, homoeologous recombination between S. acaule and S. tuberosum chromosomes is possible and useful traits from S. acaule may be transferred to the S. tuberosum gene pool. This revised version was published online in July 2006 with corrections to the Cover Date.     

Development of SCAR markers linked to the Ns locus in potato

A random amplified polymorphic DNA marker OPG17₄₅₀ linked to the Ns gene that confers resistance of potato to potato virus S (PVS), was used to develop sequence‐characterized amplified region (SCAR) markers. After cloning and sequencing of OPG17₄₅₀ new polymerase chain reaction (PCR) primers were designed to generate dominant (SCG17₃₂₁) and codominant (SCG17₄₄₈) markers. For SCG17₄₄₈, polymorphism between susceptible and resistant genotypes was recovered after digestion of the marker with the restriction enzyme Muni. In addition to the band corresponding to ‘susceptible’ allele that does not contain the Muni cleavage site, two bands of approximately 251 bp and 197 bp were observed in the resistant genotypes. The usefulness of these SCAR markers was verified in diploid potatoes possessing the Ns locus from clone G‐LKS 678147/60, and in tetraploid potatoes derived from G‐LKS 678147/60 and from clone MPI 65118/3.     

Localization of the Laevigatum Resistance Gene MlLa against Powdery Mildew in the Barley Genome by the Use of RFLP Markers

RFLP markers which were previously assigned to chromosome 2 (2H) were found to detect polymorphism between the cv. ‘Pallas’ and a near isogenic line carrying the Laevigatum resistance gene MlLa. Linkage analysis carried out with two sets of DH lines derived from the crosses ‘RisøS’בSultan’ and ‘Alf’בVogelsanger Gold 2’ confirmed three DNA probes closely linked with the MlLa locus.     

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.     

Monohaploid plants from anthers of a dihaploid genotype of Solanum tuberosum L.

Summary Monohaploid Solanum tuberosum plants were produced from the anthers of a dihaploid genotype. From another dihaploid genotype plants containing 36 chromosomes were obtained. For plantlet production anthers containing pollen at the uninucleate microspore stage were inoculated on a Linsmaier and Skoog-based medium supplemented with 1 mg/1 indole-3-acetic acid and 1 mg/1 benzyl aminopurine.Anthers from donor plants grown either in the summer or in the winter responded similarly on a range of media. Anther response in the form of callus induction and root formation was at least partly dependent on the genotype of the donor dihaploid plant.     

应用双单倍体克服籼粳交的不亲和性初探

通过对籼粳交Ｆ１体细胞愈伤组织的秋水仙素处理和四倍体花药培养，建成了水稻四倍体诱导及双单倍体分解系统。应用这一系统获得了４４个籼粳交组合的双单倍体及其自交４后代。对这些双单倍体及其后代的遗传分析表明，籼粳交育性可能受寡基因控制，双单倍体途径通过改变正常的世代交替过程，能部分克服籼粳交的不育性及倾籼或倾粳的偏态分离，并具有比Ｆ２更丰富的遗传多样性。在双单倍体一代群体中，育性正常的植株约占４０％，这是     

Genetics of self-compatibility in dihaploids of Solanum tuberosum L. 4. Linkage between an S-bearing translocation and a locus for virescens

Summary Three dihaploids of Solanum tuberosum (two self-compatible, one self-incompatible) were found to be heterozygous for a monogenic recessive virescent mutant. Intercrossing resulted in the expected 3 : 1 ratio only in crosses involving one self-compatible and one self-incompatible parent. Self-compatible x self-compatible matings produced F₁'s in which 6:1 was found. The same ratio was observed in the self progeny of the two self-compatible dihaploids. This significant deviation could be explained by assuming linkage (25% crossing-over) between v and an S-bearing translocation. This translocation causes self-compatibility in the dihaploids used and early lethality when homozygous.     

Female fertility of potato (Solanum tuberosum ssp. tuberosum) dihaploids

Summary Results of testcrosses and seed set data of the dihaploid breeding program at the Potato Research Institute in Gross Lüsewitz, Germany, indicate that the low level of female fertility could hinder success in crossing work. With the objective to reveal major components of female fertility, the behavior of 47 outstanding potato dihaploids (S. tuberosum ssp. tuberosum, 2n=2x=24) was assessed in testcrosses over a period of 3 years. Analysis of variance was carried out for the traits: number of seed per berry (S/B), number of intact seed per berry (IS/B), and number of non-germinable seed per berry (NGS/B). Significant sources of variation included cross combinations, years, and cross combination-year interactions. More than 20 S/B were obtained in 25.4% of all cross combinations performed over 3 years using up to three pollinators. Berry set was not correlated to seed set, but was in general determined by the pollinator used. The number of seedless berries (SLB/F) was related to neither total berry set nor seed production. The trait S/B remained stable over different years, and thus it deemed to be most useful as a measure of the level of female fertility in practical breeding work. Selection for female fertility in early generations of a breeding program is recommended to avoid limitations in the production of variability via sexual recombination.