Hi Matt, great software! I had a couple of questions regarding the methylation calling settings and the modkit usage you detail in the ROBIN paper.
In MinKNOW, there's an option to select (wording might not be exact) "5hmC/5mC in all contexts". My understanding is that this setting is primarily for non-mammalian species where there are both CpG and CHG (H = A/T/C) islands contributing to methylation. In your opinion, would having the all contexts setting enabled in MinKNOW be detrimental to the analysis, potentially calling methylation at 'false' CpG sites?
Secondly, in the ROBIN paper, you run “modkit” with “adjust-mods --convert h m” to convert 5hmC calls to 5mC calls in the output bam files. I wondered if this was now done automatically by ROBIN or if it's a standalone script pre-ROBIN? There is code pertaining to COMBINED_MOD_CODE in the matkit.py utility but I don't think this achieves the same thing as modkit. If modkit is run before ROBIN, how does this operate in tandem with real-time ROBIN runs, i.e. how do you stream the output bams from MinKNOW -> modkit -> ROBIN?
Thanks in advance, Oliver
Hi Matt, great software! I had a couple of questions regarding the methylation calling settings and the modkit usage you detail in the ROBIN paper.
In MinKNOW, there's an option to select (wording might not be exact) "5hmC/5mC in all contexts". My understanding is that this setting is primarily for non-mammalian species where there are both CpG and CHG (H = A/T/C) islands contributing to methylation. In your opinion, would having the all contexts setting enabled in MinKNOW be detrimental to the analysis, potentially calling methylation at 'false' CpG sites?
Secondly, in the ROBIN paper, you run “modkit” with “adjust-mods --convert h m” to convert 5hmC calls to 5mC calls in the output bam files. I wondered if this was now done automatically by ROBIN or if it's a standalone script pre-ROBIN? There is code pertaining to COMBINED_MOD_CODE in the matkit.py utility but I don't think this achieves the same thing as modkit. If modkit is run before ROBIN, how does this operate in tandem with real-time ROBIN runs, i.e. how do you stream the output bams from MinKNOW -> modkit -> ROBIN?
Thanks in advance, Oliver