Variant filtering
STU-66 family: ES data of the four members, resulted in 73,426
variants in affected father, 73,689 in unaffected mother, 73,141 in
unaffected sibling and 73,198 variants in proband.
Applying variant filter criteria (detailed in methodology), resulted in
a mean of 2245 variants (ranging from 2188-2296). The variant data of
the four individuals were analyzed using Venn diagram that produced 15
possible combinations of the data (figure 3). One such combination
revealed 276 genes that were common to affected father and the proband
(figure A1). On filtering further for the common variants in them ensued
260 variants harbored in 248 genes. Of these 260 common variants, 218
were previously known (dbSNP database) and 42 were novel variants
(hitherto unreported). Only 85 of the 218 known variants and 12 of the
42 novel variants fall in the coding region. Applying a four step
exclusion criteria (detailed in methodology) to known variants,
shortened the list to 18 genes (all heterozygous variants). The genes of
interest thus sums up to 19 known variants and 12 novel variants, spread
across 30 genes (table A1).
STU-65 family: ES data of the two affected sibs (V-33, proband
and V-35, affected brother) resulted in 58,346 and 51,062 variants
respectively. Applying similar criteria as described above using Venn
diagram approach, 294 genes were found common to the sib-pair (figure
A2). The downstream analysis of the variants observed in the 294 common
genes is depicted in the flow chart (figure A3). Those variants common
to both sibs were sorted out resulting in 161 common variants harbored
in 153 genes. Of these, 158 were known variants and 3 were novel
variants that were all found in the coding regions. Applying the same
four step exclusion criteria (as in STU-66) to 158 known variants,
narrowed down the gene list to 34 variants (4 homozygous and 30
heterozygous). Thus the genes of interest sum up to 31 known and 3 novel
variants, in 34 genes. (table A2).
No significant intronic/UTR variation was detected, other than the
indels and missense variations, in the exome data of both families.
Considering stuttering to be a less common trait, an allele count of
more than 20 in gnomAD database were excluded, narrowing the list to 35
genes.