A sequence reference library is not a prerequisite, but it
is something better than nothing
Soil sample source tracking using DNA metabarcoding (or any other
methods) is based on a set of data (here considered a local library) and
operation taxonomic units (OTUs) instead of species names are used. This
means that a universal reference sequence library is not necessary for
forensics. As exemplified in this study, results based on the total OTUs
came to the same conclusion as that based on the annotated OTUs.
However, if the OTUs were annotated to species, genera or families,
extra information such as morphological characters could be used and the
evidences would be more solid.
To annotate the OTUs, a well-curated sequence reference library is
indispensable. The reference library helps to exclude data of
experimental artifacts (such as chimeras) and non-target species.
Although some efforts have been made, the DNA barcode reference library
is still far from being satisfactory due to low species coverage (Lou et
al., 2010; Ratnasingham & Hebert, 2007; Tnah et al., 2019), especially
for lower plants. For example, there are about 8397 known species
worldwide and only 889 species have their rbcL sequences
deposited in GenBank (4116 accessions. accessed on Jan. 9, 2021,rbcL in plastid genomes were not considered.).