Unknown,Transcriptomics,Genomics,Proteomics

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Dominance hierarchy arising from the evolution of a complex small RNA regulatory network

ABSTRACT: Self-pollen rejection in the Brassicaceae is determined by the diploid genotype of the pollen-producing plant, and it has long been known that the alleles show a dominance hierarchy. How this hierarchy is controlled and evolves has been a classical puzzle since the pre-molecular days of genetics. Here, we uncover the system of at least 17 small RNA (sRNA) producing loci and their multiple target sites that collectively control the dominance hierarchy among alleles of a self-incompatible Arabidopsis species. Our results demonstrate that natural selection shapes a dynamic repertoire of sRNA/targets interactions by jointly acting on sRNA genes, their processing precision and their target sites. It is remarkable that a single gene can evolve such a complex system of regulation among its own alleles. Sequencing of small RNAs from A. halleri floral buds, with this single exception (Al14, from A. lyrata).

ORGANISM(S): Arabidopsis halleri

SUBMITTER: Vincent Castric

PROVIDER: E-GEOD-57267 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Targeted genomic enrichment followed by next-generation sequencing dramatically increased the efficiency of mutation discovery in human genomes. Here we demonstrate that these techniques also revolutionize traditional genetic approaches in model systems. We developed a two-step protocol utilizing a traditional bulk-segregant analysis (BSA) approach for positional cloning mutants in phenotype-driven forward genetic screens. First, BSA pools are 'light' sequenced for rough mapping, followed by targeted enrichment and deep-sequencing of the mutant BSA pool for the linked genomic region to fine-map and discover candidate mutations. We applied this method successfully to three Arabidopsis mutants and show that it can be scaled by multiplexing. Similarly, we applied these techniques to a gene-driven reverse genetics method (chemical driven target-selected mutagenesis or TILLING) that is used for generating gene knockouts in a wide range of organisms, including plants, invertebrates and vertebrates. We developed an efficient multiplexed genomic enrichment protocol for pre-barcoded samples. As a proof-of-principle, 770 genes were screened for induced mutations in 30 rats, which identified all but one known variants (30) as well as a large series of novel knockout and missense alleles. Mutations were retrieved at the expected frequency with a the false-positive rate of less than 1 in 6 million basepairs, which is much lower as compared to traditional mutation discovery approaches. Both methods are largely independent of the genome size due to the targeted enrichment and can thus be applied to any genetic model system of interest. Targeted genomic enrichment followed by next-generation sequencing dramatically increased the efficiency of mutation discovery in human genomes. Here we demonstrate that these techniques also revolutionize traditional genetic approaches in model systems. We developed a two-step protocol utilizing a traditional bulk-segregant analysis (BSA) approach for positional cloning mutants in phenotype-driven forward genetic screens. First, BSA pools are 'light' sequenced for rough mapping, followed by targeted enrichment and deep-sequencing of the mutant BSA pool for the linked genomic region to fine-map and discover candidate mutations. We applied this method successfully to three Arabidopsis mutants and show that it can be scaled by multiplexing. Similarly, we applied these techniques to a gene-driven reverse genetics method (chemical driven target-selected mutagenesis or TILLING) that is used for generating gene knockouts in a wide range of organisms, including plants, invertebrates and vertebrates. We developed an efficient multiplexed genomic enrichment protocol for pre-barcoded samples. As a proof-of-principle, 770 genes were screened for induced mutations in 30 rats, which identified all but one known variants (30) as well as a large series of novel knockout and missense alleles. Mutations were retrieved at the expected frequency with a the false-positive rate of less than 1 in 6 million basepairs, which is much lower as compared to traditional mutation discovery approaches. Both methods are largely independent of the genome size due to the targeted enrichment and can thus be applied to any genetic model system of interest.

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Dominance hierarchy arising from the evolution of a complex small RNA regulatory network

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