A pilot experiment with known SNPs showed that they could be detected at a frequency > 0.3% within the pools. Barcodes were incorporated during PCR, and the pooled amplicons were sequenced using an Ion Torrent PGM. We used an amplicon-based approach in which DNA samples from an ethyl methanesulfonate (EMS)-mutagenized population were pooled and used as template in PCR reactions to amplify a region of each gene of interest. We developed this method in flax ( Linum usitatissimum), to demonstrate its utility in a crop species. As an alternative to these genome or exome-scale methods, we sought to develop a scalable and efficient method for detection of induced mutations that could be applied to a small number of target genes, using Ion Torrent technology. exome capture, whole genome resequencing). Many reverse genetics approaches have been developed to identify mutations in genes of interest within a mutagenized population, including some approaches that rely on next-generation sequencing (e.g. Detection of induced mutations is valuable for inferring gene function and for developing novel germplasm for crop improvement.
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