Project description:DNA sequencing using nanopore technologies with the affordable MinION device is useful for the identification and characterization of structural variants, long haplotypes, sequencing of repetitive regions and identification of epigenetic modifications. The main limitation of this approach is the low coverage obtained, which might be avoided by adaptive sampling, a computationally controlled method of enrichment for targeted genomic regions. This study dissects the factors involved in the enrichment by adaptive sampling of a panel of 18 human genome regions containing 20 genes implicated in breast cancer sequenced in 16 patients with familial breast cancer negative for NGS screening. An average coverage of 2.0x was obtained for the whole genome and 5.1x for selected regions. Sequencing time was the main factor improving coverage. The selection of long reads (>1 Kb) did not improve the enrichment. The length of the selected region, which in our study ranged from 126 to 565 Kb, did not play a significant role in enrichment. However, the region containing PMS2 showed significantly lower coverage, which could be explained by the high number of PMS2 pseudogenes (N = 14), which were also enriched. Our study shows new evidence of enrichment obtained by adaptive sampling in a panel of genomic regions and shows parameters, the relevance of sequencing time and the role of pseudogenes, that improve the enrichment yield with no library reloading or GPU use, data useful for a more efficient application of this procedure in future studies.
Project description:CDCA7-associated global aberrant DNA hypomethylation translates to localized, tissue-specific transcriptional responses [ONT adaptive sampling]
Project description:We have developed an endovascular catheter-based device for sampling of the endothelium, intended for RNA-seq analysis. With an outer diameter of 0.23 mm it is capable of reaching peripheral vessels for endothelial sampling, and favourably compares to other approaches with stent retrievers or similar tools. Here, we have collected samples from liver and kidney vessels from swine and performed RNA-sequencing to validate the sample quality.
