Hi,
I plan to conduct detailed research on the endophytic microbiome in one of Apiaceae members. I will collect root, leaf and seed samples to study endophytic microbiome. I will also analyse soil microbiota as a reference. I will perform 16S rRNA and ITS metabarcoding (so both Bacteria and Fungi). I'm not sure what homogenisation & extraction method I should use. I've done some research, but I'm still unsure. When it comes to the plant tissues there is no clear recommendation, and when talking about "various sets of samples for metagenomics", authors do not typically include plant endomicrobiome.
So my question is: should I (of course after sterilisation) homogenise the plant tissues in liquid nitrogen to powder? And then? What's your experience? Should I perform SDS-based or CTAB-based DNA extraction or use some commercial kits? If the latter is the solution, is there an alternative to the PowerSoil kit? The research grant budget for PhD candidates in my country is quite limited, so PowerSoil (Qiagen) is too expensive.
Or maybe I should combine some methods?
I know the cell lysis stage is critical; in the next steps I will also use an internal synthetic spike-in standard (to perform quantitative NGS), but this method assumes 100% cell lysis efficiency, that's why it is so important to me.
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