Figure 1
Schematic representation of the metagenomic approach. Total genomic DNA was extracted from dried leaf material, followed by a rolling circle amplification (RCA). A random amplification step using polymerase chain reaction (RA-PCR) combined with a tagging was performed. The use of distinct tags, here symbolized with the «T1» and «T2» labels, allows each sequence to be traced back to the original sample. During the multiplexing step, amplicons are equimolarly pooled before being submitted to Illumina library construction and sequencing. Mastrevirus sequences identified using similarity search (sequences with green ticks on the figure) were more precisely classified through phylogenetic placement analysis on mastreviruses reference alignment and tree (in grey). Viral classifications are then confirmed after cloning and Sanger sequencing using the RCA-RFLP protocol.
