Project description:We performed direct cDNA sequencing in HeLa GFP∆Promoter cells by Oxford Nanopore Technology (ONT) on a MinION device to detect EGFP RNA levels after DSB induction.
Project description:Whole Genome Bisulfite Sequencing (WGBS) has been the gold standard DNA methylation mapping and quantification for over a decade. Oxford Nanopore Technologies (ONT) sequencing directly measures nucleotide modifications. In this study, we have compared DNA methylation levels (5-methylcytosine) at CpG sites in the quail genome using WGBS and ONT. Samples were collected to investigate transgenerational DNA methylation changes in Japanese quail following ancestral exposure to a phytoestrogen. Blood samples from 24 third-generation (G3) individuals—descendants of either treated or untreated ancestors—were sequenced after bisulfite conversion. Both methods revealed broadly consistent methylation patterns. ONT reads covered more CpG sites and detected a higher number of differentially methylated cytosines (DMCs). Principal component analyses showed that both sex and ancestral treatment groups accounted for a portion of the observed epigenetic variation, for both technologies. Strong concordance between WGBS and ONT results supports the reliability of ONT sequencing for epigenomic research, including in quails. These data pave the way for further investigation into whether genistein induces epigenetic changes for several generations.
Project description:Higher-order chromatin structure arises from the combinatorial physical interactions of many genomic loci. To investigate this aspect of genome architecture we developed Pore-C, which couples chromatin conformation capture with Oxford Nanopore Technologies (ONT) long reads to directly sequence multi-way chromatin contacts without amplification.
Project description:Zebrafish is a widely used model organism for investigating human diseases, including hematopoietic disorders. However, a comprehensive methylation baseline for zebrafish primary hematopoietic organ, the kidney marrow (KM), is still lacking. We employed Oxford Nanopore Technologies (ONT) sequencing to profile DNA methylation in zebrafish KM by generating four KM datasets, with two groups based on the presence or absence of red blood cells. Our findings revealed that blood contamination in the KM samples reduced read quality and altered methylation patterns. Compared with whole-genome bisulfite sequencing (WGBS), the ONT-based methylation profiling can cover more CpG sites (92.4% vs 70%-80%), and exhibit less GC bias with more even genomic coverage. And the ONT methylation calling results showed a high correlation with WGBS results when using shared sites. This study establishes a comprehensive methylation profile for zebrafish KM, paving the way for further investigations into epigenetic regulation and the development of targeted therapies for hematopoietic disorders.