Abstract
BackgroundWhite clover (Trifolium repens L.) is an outbreeding allotetraploid species and an important forage legume in temperate grassland agriculture. Comparison of sub-genome architecture and study of nucleotide sequence diversity within allopolyploids provides insight into evolutionary divergence mechanisms, and is also necessary for the development of whole-genome sequencing strategies. This study aimed to evaluate the degree of divergence between the O and P' sub-genomes of white clover through sequencing of BAC clones containing paired homoeoloci. The microsyntenic relationships between the genomes of white clover and the model legumes Lotus japonicus and Medicago truncatula as well as Arabidopsis thaliana were also characterised.ResultsA total of four paired homoeologous BACs were selected and sequenced to generate 173 kb of overlapping sequence between the O and P' sub-genomes. Equivalent gene content was generally observed, apart from small-scale deletions, in contrast to conservation of intergenic sequences, which varied between the four selected regions. Measurement of the number of synonymous substitutions between homoeologous genes led to estimation of a 4.2 million year divergence time between the two sub-genomes. Microsynteny was observed between the genomes of white clover and L. japonicus for all four targeted regions, but corresponding M. truncatula genomic regions were only identified for two BAC pairs.ConclusionsThis study describes the first analysis of sub-genome structural conservation across selected genomic regions in white clover. Although the high levels of sequence conservation between the O and P' sub-genomes would complicate efforts for whole genome sequence assembly, the conserved microsynteny with model legume genomes, especially that of L. japonicus, will be highly valuable for the future of white clover genomics and molecular breeding.
Highlights
White clover (Trifolium repens L.) is an outbreeding allotetraploid species and an important forage legume in temperate grassland agriculture
This study describes the analysis of four paired white clover homoeologous bacterial artificial chromosome (BAC) and provides the first evaluation of divergence between the co-resident O and P' subgenomes for both genic and intergenic regions
For those BACs not sequenced to completion, all analysis was performed using that contig which contained the selected gene (ZPT2, DREB3, DHNb or ANR)
Summary
White clover (Trifolium repens L.) is an outbreeding allotetraploid species and an important forage legume in temperate grassland agriculture. Comparison of sub-genome architecture and study of nucleotide sequence diversity within allopolyploids provides insight into evolutionary divergence mechanisms, and is necessary for the development of whole-genome sequencing strategies. This study aimed to evaluate the degree of divergence between the O and P' sub-genomes of white clover through sequencing of BAC clones containing paired homoeoloci. This process is highly complicated by polyploidy, especially when sub-genomes are closely related, which might be expected to dispose towards chimaeric contig assembly. Studies of homoeologous sub-genome conservation within various paleopolyploids have revealed both low (maize [7]) and high (soybean [8]) levels of gene retention, while true polyploids such as cotton contain both conserved genic and intergenic regions [9,10]
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