Cannabis is one of the most popular drugs consumed in the United States. While work has been done to quantify whole genome transgenerational effects of cannabis exposure, there has been little done to quantify transgenerational effects on a gene-by-gene basis. Previously, Reduced Representation Bisulfite Sequencing identified Shank1 and Dlg4, genes associated with the post-synaptic density and the post synaptic membrane, as being differentially methylated in the sperm profile of cannabis(THC) exposed rats and controls. It is however unknown that this difference in methylation is significant under specific gene methylation analysis. It is also unknown if any differences in methylation cause significant changes in the expression of Shank1 and Dlg4. That is why in this study, potential differential methylation of Shank1 and Dlg4* were measured through pyrosequencing of the somatic tissues of F1 rats. In addition, RT-PCR** was conducted in order to quantify changes in gene expression of somatic tissues in these F1 rats. It was found that in Shank1 there was hypermethylation in both CpG sites of interests, while in Dlg4 there was hypomethylation in one CpG site and hypermethylation in the other.*** The next steps for this study would be to quantify the gene expression and methylation changes for the F2 generation, while also quantifying the phenotype in the F1 generation.
*Biotin labeled primer for this gene(necessary for pyrosequencing) may not arrive due to inconsistencies with Dlg4’s primers in PCR
** RT-PCR may not be done at the time of this poster presentation
*** This data is from the RRBS whole genome methylation, not from specific gene methylation from pyrosequencing