The Training School on Methodologies to study root-microbe interactions offered participants a unique opportunity to combine theory with hands-on practice. From isolating and modifying microbes to applying cutting-edge molecular and imaging tools, the course provided essential skills for exploring root–microbe interactions.
Session 1
Sample harvesting: pot and field experiments
Microbe isolation (culturomics) and screens for function: rhizospheric / epiphytic / endophytic / hyphospheric microbes, dilution on solid or liquid medium, in vitro cultivable bacteria and fungi, AMF (in planta and vitro monosporal inoculation) – (geno)typing/affiliation – functional assignation (in vitro screen (-N, -P, -Fe, antibiosis, hormones…), database, genome sequencing and metabolic network reconstruction.
Session 3
Genetic modification of microbes (bacteria) introduction of reporter genes (fluorescent or enzymatic tag, pro GUS, pro GFP…): chromosome insertion vs. plasmid, bombardment, heat shock, electroporation, conjugation; loss of function mutants: recombination, CRISPR-CAS9, transposon insertions…
Microscopy: bright field – fluorescent – confocal – electronic. Staining of microorganisms,generation of reporter lines, immunolabeling
DNA and RNA extractions for metabarcoding, (meta)genomics, (meta)transcriptomics, RNA-seq, qRT-PCR. Short vs. long sequencing.
Session 6
Microbe quantification during colonization: cfu, biomass, qPCR, lipid analysis, respiration, glomalin dosage; microscopy, nodule number, nodule biomass, nitrogen fixation measurement methods (ARA and ANA)
