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Microspore culture reveals complex meiotic behaviour in a trigenomic Brassica hybrid.

Mason AS, Takahira J, Atri C, Samans B, Hayward A, Cowling WA, Batley J, Nelson MN - BMC Plant Biol. (2015)

Bottom Line: The MD progeny were also compared to progeny derived by self-pollination and open-pollination for fertility (estimated by self-pollinated seed set and pollen viability) and DNA ploidy (measured by flow cytometry).Average fertility was significantly lower in progeny obtained by microspore culture than progeny obtained by self-pollination or open-pollination, after excluding MD progeny which had not undergone chromosome doubling.SNP allele copy number analysis revealed the occurrence not only of homoeologous duplication/deletion events but also other cryptic duplications and deletions that may have been the result of mitotic instability.

View Article: PubMed Central - PubMed

Affiliation: School of Agriculture and Food Sciences, The University of Queensland, Brisbane, 4072, Australia. annaliese.mason@agrar.uni-giessen.de.

ABSTRACT

Background: Development of synthetic allohexaploid Brassica (2n = AABBCC) would be beneficial for agriculture, as allelic contributions from three genomes could increase hybrid vigour and broaden adaptation. Microspore culture of a near-allohexaploid hybrid derived from the cross (B. napus × B. carinata) × B. juncea was undertaken in order to assess the frequency and distribution of homologous and homoeologous crossovers in this trigenomic hybrid. SNP and SSR molecular markers were used to detect inheritance of A, B and C genome alleles in microspore-derived (MD) progeny. SNP allele copy number was also assessed. The MD progeny were also compared to progeny derived by self-pollination and open-pollination for fertility (estimated by self-pollinated seed set and pollen viability) and DNA ploidy (measured by flow cytometry).

Results: In the trigenomic hybrid, homologous chromosome pairs A(j)-A(n), B(j)-B(c) and C(n)-C(c) had similar meiotic crossover frequencies and segregation to that previously observed in established Brassica species, as demonstrated by marker haplotype analysis of the MD population. Homoeologous pairing between chromosomes A1-C1, A2-C2 and A7-C6 was detected at frequencies of 12-18 %, with other homoeologous chromosome regions associating from 8 % (A3-C3) to 0-1 % (A8-C8, A8-C9) of the time. Copy number analysis revealed eight instances of additional chromosomes and 20 instances of chromosomes present in one copy in somatically doubled MD progeny. Presence of chromosome A6 was positively correlated with self-pollinated seed set and pollen viability in the MD population. Many MD progeny were unable to produce self-pollinated seed (76 %) or viable pollen (53 %), although one MD plant produced 198 self-pollinated seeds. Average fertility was significantly lower in progeny obtained by microspore culture than progeny obtained by self-pollination or open-pollination, after excluding MD progeny which had not undergone chromosome doubling.

Conclusions: Based on SNP data analysis of the microspore-derived progeny, crossover frequency per chromosome in the allohexaploid hybrid was found to be similar to that in established Brassica species, suggesting that the higher chromosome number did not significantly disrupt cellular regulation of meiosis. SNP allele copy number analysis revealed the occurrence not only of homoeologous duplication/deletion events but also other cryptic duplications and deletions that may have been the result of mitotic instability. Microspore culture simplified the assessment of chromosome behaviour in the allohexaploid hybrid but yielded progeny with lower fertility and a greater range of ploidy levels compared to progeny obtained by self- or open-pollination.

No MeSH data available.


Related in: MedlinePlus

DNA ploidy level as estimated by flow cytometry in three populations derived from microspore culture, self-pollination and open-pollination of the same trigenomic hexaploid hybrid resulting from the cross (B. napus × B. carinata) × B. juncea. Only progeny with consistent DNA content between multiple samples are shown. The Genetics Society of America has granted permission for re-publication of the data from the self-pollinated population for the purposes of comparison: original data published in Mason et al. [25]
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Fig6: DNA ploidy level as estimated by flow cytometry in three populations derived from microspore culture, self-pollination and open-pollination of the same trigenomic hexaploid hybrid resulting from the cross (B. napus × B. carinata) × B. juncea. Only progeny with consistent DNA content between multiple samples are shown. The Genetics Society of America has granted permission for re-publication of the data from the self-pollinated population for the purposes of comparison: original data published in Mason et al. [25]

Mentions: DNA ploidy levels in the MD population were inferred from relative DNA content, and ranged from 0.26 to 1.0 of the expected DNA content for a 2n = AABBCC = 54 allohexaploid (Fig. 6). MD progeny with consistent ploidy results fell into two groups: putatively doubled chromosome number (hereafter referred to as “2n”) (53 plants including the unreduced gamete-derived progeny, ploidy range (relative DNA content) 0.76–1.0 and undoubled chromosome number (hereafter referred to as “n”) (6 plants, ploidy range 0.26–0.46) (Fig. 6). Of the 41 MD progeny which were sampled 2–4 times, 12 progeny had > 40 % variation between readings, suggestive of chimeric tissue (both “2n” and “n” chromosome number in sectors of the same plant) (Additional file 5: Table S4). The percentage variation between the maximum and minimum readings for each of the B. rapa, B. napus and B. carinata samples was 2–4 % (Additional file 5: Table S4). However, while some MD, SP and OP plants fell into the 0–5 % variation range, variation within the 5–37 % range was also observed for some plants and in the near-allohexaploid hybrid controls (Additional file 5: Table S4).Fig. 6


Microspore culture reveals complex meiotic behaviour in a trigenomic Brassica hybrid.

Mason AS, Takahira J, Atri C, Samans B, Hayward A, Cowling WA, Batley J, Nelson MN - BMC Plant Biol. (2015)

DNA ploidy level as estimated by flow cytometry in three populations derived from microspore culture, self-pollination and open-pollination of the same trigenomic hexaploid hybrid resulting from the cross (B. napus × B. carinata) × B. juncea. Only progeny with consistent DNA content between multiple samples are shown. The Genetics Society of America has granted permission for re-publication of the data from the self-pollinated population for the purposes of comparison: original data published in Mason et al. [25]
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4493989&req=5

Fig6: DNA ploidy level as estimated by flow cytometry in three populations derived from microspore culture, self-pollination and open-pollination of the same trigenomic hexaploid hybrid resulting from the cross (B. napus × B. carinata) × B. juncea. Only progeny with consistent DNA content between multiple samples are shown. The Genetics Society of America has granted permission for re-publication of the data from the self-pollinated population for the purposes of comparison: original data published in Mason et al. [25]
Mentions: DNA ploidy levels in the MD population were inferred from relative DNA content, and ranged from 0.26 to 1.0 of the expected DNA content for a 2n = AABBCC = 54 allohexaploid (Fig. 6). MD progeny with consistent ploidy results fell into two groups: putatively doubled chromosome number (hereafter referred to as “2n”) (53 plants including the unreduced gamete-derived progeny, ploidy range (relative DNA content) 0.76–1.0 and undoubled chromosome number (hereafter referred to as “n”) (6 plants, ploidy range 0.26–0.46) (Fig. 6). Of the 41 MD progeny which were sampled 2–4 times, 12 progeny had > 40 % variation between readings, suggestive of chimeric tissue (both “2n” and “n” chromosome number in sectors of the same plant) (Additional file 5: Table S4). The percentage variation between the maximum and minimum readings for each of the B. rapa, B. napus and B. carinata samples was 2–4 % (Additional file 5: Table S4). However, while some MD, SP and OP plants fell into the 0–5 % variation range, variation within the 5–37 % range was also observed for some plants and in the near-allohexaploid hybrid controls (Additional file 5: Table S4).Fig. 6

Bottom Line: The MD progeny were also compared to progeny derived by self-pollination and open-pollination for fertility (estimated by self-pollinated seed set and pollen viability) and DNA ploidy (measured by flow cytometry).Average fertility was significantly lower in progeny obtained by microspore culture than progeny obtained by self-pollination or open-pollination, after excluding MD progeny which had not undergone chromosome doubling.SNP allele copy number analysis revealed the occurrence not only of homoeologous duplication/deletion events but also other cryptic duplications and deletions that may have been the result of mitotic instability.

View Article: PubMed Central - PubMed

Affiliation: School of Agriculture and Food Sciences, The University of Queensland, Brisbane, 4072, Australia. annaliese.mason@agrar.uni-giessen.de.

ABSTRACT

Background: Development of synthetic allohexaploid Brassica (2n = AABBCC) would be beneficial for agriculture, as allelic contributions from three genomes could increase hybrid vigour and broaden adaptation. Microspore culture of a near-allohexaploid hybrid derived from the cross (B. napus × B. carinata) × B. juncea was undertaken in order to assess the frequency and distribution of homologous and homoeologous crossovers in this trigenomic hybrid. SNP and SSR molecular markers were used to detect inheritance of A, B and C genome alleles in microspore-derived (MD) progeny. SNP allele copy number was also assessed. The MD progeny were also compared to progeny derived by self-pollination and open-pollination for fertility (estimated by self-pollinated seed set and pollen viability) and DNA ploidy (measured by flow cytometry).

Results: In the trigenomic hybrid, homologous chromosome pairs A(j)-A(n), B(j)-B(c) and C(n)-C(c) had similar meiotic crossover frequencies and segregation to that previously observed in established Brassica species, as demonstrated by marker haplotype analysis of the MD population. Homoeologous pairing between chromosomes A1-C1, A2-C2 and A7-C6 was detected at frequencies of 12-18 %, with other homoeologous chromosome regions associating from 8 % (A3-C3) to 0-1 % (A8-C8, A8-C9) of the time. Copy number analysis revealed eight instances of additional chromosomes and 20 instances of chromosomes present in one copy in somatically doubled MD progeny. Presence of chromosome A6 was positively correlated with self-pollinated seed set and pollen viability in the MD population. Many MD progeny were unable to produce self-pollinated seed (76 %) or viable pollen (53 %), although one MD plant produced 198 self-pollinated seeds. Average fertility was significantly lower in progeny obtained by microspore culture than progeny obtained by self-pollination or open-pollination, after excluding MD progeny which had not undergone chromosome doubling.

Conclusions: Based on SNP data analysis of the microspore-derived progeny, crossover frequency per chromosome in the allohexaploid hybrid was found to be similar to that in established Brassica species, suggesting that the higher chromosome number did not significantly disrupt cellular regulation of meiosis. SNP allele copy number analysis revealed the occurrence not only of homoeologous duplication/deletion events but also other cryptic duplications and deletions that may have been the result of mitotic instability. Microspore culture simplified the assessment of chromosome behaviour in the allohexaploid hybrid but yielded progeny with lower fertility and a greater range of ploidy levels compared to progeny obtained by self- or open-pollination.

No MeSH data available.


Related in: MedlinePlus