Sequencing of actual transcribed sequences of THCAS and CBDAS from RNA
obtained from fully flowering Cannabis inflorescences, partially reduced the
complexity of the cannabinoid synthase sequences found. In a recent work, with
this approach we obtained nine THCAS and twelve CBDAS sequences upon
sequencing 18 inbred lines with different chemotypes (I, III and IV); variability in
the number of the main transcribed sequences was found, as from one up to 5
different synthases were found expressed in the inflorescences (Onofri et al. 2015 ).
Sequences with specific SNPs, putatively leading to the production of cannabinoid
synthases with different and/or reduced efficiencies were identified, further
enriching the repertoire of these enzymes, but on the other hand making devel-
opment of chemotype-specific markers an increasingly difficult task.
Similar results were also obtained by sequencing the chemotype-related genes in
different THCA-prevalent, CBDA-prevalent and their hybrids, confirming once
more the sequence complexity already described in both putatively functional and
nonfunctional cannabinoid synthase-related pseudogenes (McKernan et al. 2015 ).
In the light of what has been described above, the mandatory importance of taking
into consideration the increasing number of SNPs discovered in future
chemotype-associated marker design becomes clear.
The high number of sequences of the cannabinoid synthase genes available
reflects the high degree of variation present in theCannabisgenomes, and points to
the existence of a true gene family. A cluster analysis of all the sequences available
clearly separated CBDAS from THCAS sequences (Onofri et al. 2015 ); Fig.15.5
illustrates the relationships occurring between a number of sequences taken from
the databases. Interestingly, a“transition”group formed by sequences AB292683
and AB292684 (Taura et al. 2007 ), putatively non-functional CBDAS sequences,
and two functional THCAS identified in a drug and in afiber strain by Weiblen
et al. ( 2015 ), was identified, located between the two main groups.
Fig. 15.4 Distribution of CBDA/THCA ratios in two different segregating F2 populations. Note
the difference in CBDA/THCA ratio in the two different offsprings, corresponding to different
relative efficiencies of the two synthases. This difference has been shown to be fully inherited from
the respective F1 plants
336 C. Onofri and G. Mandolino