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Plant Materials: Brassica Polyploids

The genus Brassica includes three diploid species and three amphidiploid species. The diploid species probably evolved from ancient polyploids because they have triplicated copies of most genome segments. The amphidiploid species have evolved relatively recently by hybridization of the diploid species, although their exact diploid progenitors are not known. Each of the amphidiploid species can be resynthesized by crossing diploids, rescuing embryos and colchicine doubling the progenies. This provides completely homozygous genotypes (with known progenitors) that should breed true in future generations. Alternatively, one can create autotetraploids of the diploid species and cross these to create the amphidiploid. If one starts with diploids derived from double haploids, the resulting polyploids should be completely homozygous and breed true.

Genome relationships of the Brassica species. The diploid species are at the corners of the triangle, the amphidiploid species are between their diploid progenitors. Genome composition (capital letters for cytoplasmic, small letters for nuclear genomes) and haploid chromosome numbers are shown.

We have resynthesized B. napus polyploids by reciprocally crossing B. rapa (cv Flowering Pak Choi) by B. oleracea (rapid cycling genotype) followed by embryo rescue and chromosome doubling (Song et al. 1993, Theor. Appl. Genet. 86:811-821). The advanced inbreds of these resynthesized B. napus polyploids were used to study genome changes (Song et al. 1995, PNAS 92:7719-7723) and de novo variation in flowering time (Schranz and Osborn 1999, J. Hered. 91:242-246); however they are self-incompatible and difficult to advance by self-pollination. We also have observed de novo variation in flowering time among another set of genetically identical lines derived from a single resynthesized B. napus plant that was obtained from a cross of B. oleracea (rapid cycling) and B. rapa (oilseed cv Reward). These lines are self-compatible and produce high quantities of seeds. Early- and late-flowering selections from this lineage are being used in microarray experiments to test the magnitudes of sampling variation and changes in expression of flowering time genes.

Early- and late-flowering lines were selected from a resynthesized B. napus lineage. All plants were derived from a single chromosome-doubled amphihaploid and should be genetically identical. However, the parents were from open-pollinated seed stocks and are not homozygous.

A new set of 50 B. napus polyploids have been developed by crossing a rapid cycling, self-compatible doubled haploid lines of B. oleracea and B. rapa . Ten of these hybrids spontaneously doubled and the remainder were treated with colchicine. We also have created the autotetraploids of the parents. These 50 polyploids are being advanced by self-pollination of single plants for several generations. We have analyzed these in the S1 and S5 generations for changes in gene expression and genome structure. The results tell us that the effects of polyploidy are not necessarily similar across many independently derived lines. We observed changes that changes in gene expression can occur rapidly after formation. Changes are also observed in the later generations and are accompanied by changes in genome structure which are consistent with chromosomal translocation between homeologous chromosomes.

Development of new resynthesized B. napus lines from double haploid lines of B. oleracea and B. rapa . All B. napus lines should be genetically identical.
Phenotypic instabilities in new B. napus polyploids. White flower color was inherited as dominant trait from B. oleracea , but some S0 polpyloids showed sectors of yellow flowers. This was not observed in the S1 generation.

Brassica lines are available through the Arabidopsis Biological Resource Center.

TO 1000 DH3 Brassica oleracea Doubled Haploid, self-compatible, white-flowered, diploid parent of resynthesized Brassica napus allopolyploids.
IMB 218 DH1 Brassica rapa Doubled Haploid, self-compatible, yellow-flowered, diploid parent of resynthesized Brassica napus allopolyploids.
50 resynthesized Brassica napus . To be deposited after seed increase of S5 generation sets seed.

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