Introduction of beet cyst nematode resistance from Sinapis alba L. and Raphanus sativus L. into Brassica napus L. (oil-seed rape) through sexual and somatic hybridization

Experiments were performed to select for beet cyst nematode (Heterodera schachtii Schm., abbrev. BCN) resistant genotypes of Brassica napus L. (oilseed rape), and to introduce BCN-resistance from the related species Raphanus sativus L. (oil-radish) and Sinapis alba L. (white mustard) into oil-seed rape.Sexual hybridization between B.napus and R. sativus did not result in hybrid plants, whereas from about 800 crosses between B.napus and the amphidiploid xBrassicoraphanus Sageret 284 F 1 hybrid plants were obtained. Sexual hybridization between B.napus and S. alba was only successful when diploid accessions of S.alba were used as the female parent. Crossability between these species was poor; only six hybrids were obtained out of approximately 10,000 crosses. The poor crossability in the intergeneric crosses was shown to be the cause of various breeding barriers. Somatic hybridization between B.napus and either R. sativus or S. alba resulted in a few somatic hybrid plants. Putative F 1 hybrids and somatic hybrid plants were characterized by their morphology, cytology, by DNA-analysis and by scoring resistance to BCN. Somatic hybrid plants were found to be unstable for the number of chromosomes and for BCN- resistance. Some F 1 hybrids, somatic hybrids and BC 1 plants, derived from crossing F 1 hybrids to B.napus as male parent had a high level of BCN-resistance, not different from that of the resistant parental genotypes. Finally, the mechanism of resistance to BCN in resistant S. alba, R. sativus and xBrassico-raphanus was expressed in F 1 hybrids derived from crosses between resistant genotypes of these three species and B. napus.</em

Method of producing haploid and doubled haploid plant embryos

The invention relates to a method for producing haploid plant embryos, comprising providing microspores or pollen that comprise cell division inducing molecules; pollinating an embryo sac cell, in particular an egg cell, of the plant of which the haploid embryo is to be made with the microspores or pollen; allowing the microspores or pollen to discharge the cell division inducing molecules in or in the vicinity of the embryo sac cell, in particular the egg cell, to trigger division thereof to obtain a haploid plant embryo. When doubled haploid plant embryos are to be produced doubling of the chromosome number takes place at a certain stage after pollination, in particular during cell division or after obtaining the embryo. The invention further relates to the embryos thus obtained, plants regenerated therefrom and progeny thereof

Vegetative propagation of Alstroemeria hybrids in vitro.

Terminal and lateral tips from fleshy rhizomes of Alstroemeria hybrids were isolated in vitro and induced to form a new rhizome. The cultivar Toledo was used in most experiments, but later other cultivars were also tested. The basic culture medium for rhizome isolation and for rhizome multiplication was: Murashige and Skoog (MS) macro- and micro-salts at full strength (except Fe), NaFeEDTA 25 mg/l, saccharose 3°BA 2–4 mg/l vitamin B1 0.4 mg/l, and Difco Bacto-agar 0.7 &Eth;The basic culture medium for rooting was slightly different: saccharose 5°BA was omitted and 0.5 mg/l NAA was added. Rhizome cultures were placed at 21°C and 8 h fluorescent light/16 h darkness. Rooting was carried out at 21°C and 16 h fluorescent light/8 h darkness. Rhizome multiplication required a cytokinin in the medium; BA and PBA were most effective, whereas kinetin, 2iP, and zeatin were not very effective. BA at 2–4 mg/l partially suppressed erect shoot growth and stimulated rhizome branching. Addition of auxin had no effect on rhizome multiplication. Relative small rhizome explants (with one bud) had a higher multiplication rate than large ones. Optimal rhizome multiplication required 3 week cycles of subculturing; cycles of 4, 5 and 6 weeks being less productive. The multiplication rate was increased by growing the rhizomes in liquid media; however, this resulted in vitrification. Excised rhizome explants can be rooted by subculturing rhizome explants on cytokinin-free media containing auxin. Generally NAA (optimum 0.5 mg/l) induced better rooting than IBA. In vitro rooted plants were successfully transferred to the greenhouse and developed into normal flowering plants

Vegetative propagation of Alstroemeria hybrids in vitro.

Terminal and lateral tips from fleshy rhizomes of Alstroemeria hybrids were isolated in vitro and induced to form a new rhizome. The cultivar Toledo was used in most experiments, but later other cultivars were also tested. The basic culture medium for rhizome isolation and for rhizome multiplication was: Murashige and Skoog (MS) macro- and micro-salts at full strength (except Fe), NaFeEDTA 25 mg/l, saccharose 3°BA 2–4 mg/l vitamin B1 0.4 mg/l, and Difco Bacto-agar 0.7 &amp;Eth;The basic culture medium for rooting was slightly different: saccharose 5°BA was omitted and 0.5 mg/l NAA was added. Rhizome cultures were placed at 21°C and 8 h fluorescent light/16 h darkness. Rooting was carried out at 21°C and 16 h fluorescent light/8 h darkness. Rhizome multiplication required a cytokinin in the medium; BA and PBA were most effective, whereas kinetin, 2iP, and zeatin were not very effective. BA at 2–4 mg/l partially suppressed erect shoot growth and stimulated rhizome branching. Addition of auxin had no effect on rhizome multiplication. Relative small rhizome explants (with one bud) had a higher multiplication rate than large ones. Optimal rhizome multiplication required 3 week cycles of subculturing; cycles of 4, 5 and 6 weeks being less productive. The multiplication rate was increased by growing the rhizomes in liquid media; however, this resulted in vitrification. Excised rhizome explants can be rooted by subculturing rhizome explants on cytokinin-free media containing auxin. Generally NAA (optimum 0.5 mg/l) induced better rooting than IBA. In vitro rooted plants were successfully transferred to the greenhouse and developed into normal flowering plants

Vegetatieve vermeerdering van Alstroemeria in vitro.

In dit artikel worden de methoden beschreven hoe Alstroemeria efficient via in-vitrocultuur van stukjes rhizoom (wortelstok) kunnen worden vermeerderd. De methoden en adviezen zijn vooral van belang voor cultivars die zich normaal langzaam vermeerdere

Reverse breeding in Arabidopsis thaliana generates homozygous parental lines from a heterozygous plant

Traditionally, hybrid seeds are produced by crossing selected inbred lines. Here we provide a proof of concept for reverse breeding, a new approach that simplifies meiosis such that homozygous parental lines can be generated from a vigorous hybrid individual. We silenced DMC1, which encodes the meiotic recombination protein DISRUPTED MEIOTIC cDNA1, in hybrids of A. thaliana, so that non-recombined parental chromosomes segregate during meiosis. We then converted the resulting gametes into adult haploid plants, and subsequently into homozygous diploids, so that each contained half the genome of the original hybrid. From 36 homozygous lines, we selected 3 (out of 6) complementing parental pairs that allowed us to recreate the original hybrid by intercrossing. In addition, this approach resulted in a complete set of chromosome-substitution lines. Our method allows the selection of a single choice offspring from a segregating population and preservation of its heterozygous genotype by generating homozygous founder line

Hybrid recreation by reverse breeding in Arabidopsis thaliana

Hybrid crop varieties are traditionally produced by selecting and crossing parental lines to evaluate hybrid performance. Reverse breeding allows doing the opposite: selecting uncharacterized heterozygotes and generating parental lines from them. With these, the selected heterozygotes can be recreated as F1 hybrids, greatly increasing the number of hybrids that can be screened in breeding programs. Key to reverse breeding is the suppression of meiotic crossovers in a hybrid plant to ensure the transmission of nonrecombinant chromosomes to haploid gametes. These gametes are subsequently regenerated as doubled-haploid (DH) offspring. Each DH carries combinations of its parental chromosomes, and complementing pairs can be crossed to reconstitute the initial hybrid. Achiasmatic meiosis and haploid generation result in uncommon phenotypes among offspring owing to chromosome number variation. We describe how these features can be dealt with during a reverse-breeding experiment, which can be completed in six generations (~1 year