Figure legends:
Figure 1. Pedigree showing segregation of a 145 Kbp duplication
at 18q21.1, including exons 1-6 of SMAD2 . Proband is indicated
with a black arrow (D4).
Figure 2. arr[GRCh37] 18q21.1(45384387_45529362)x3 mat
duplication in proband. A) Screenshot of the duplication position as
seen in ChAS software. The duplication is shown in the first horizontal
panel as a blue rectangle. Probe positions are shown in the middle panel
as dots. Notice the elevated signal for probes directly below the blue
rectangle, which were used for calling the duplication positions. Gene
positions (SMAD2 and ZBTB7C ) are shown in the third panel,
along with the nucleotide positions in chromosome 18. Exon numbers are
displayed for SMAD2 . The shaded rectangle encompassesSMAD2 positions, and shows the partial duplication. The 145 Kbp
duplication overlapped exons one through six of SMAD2 , which
could potentially disrupt the gene’s function. In addition, the
duplication was found to be maternally inherited in proband, but the
mother was phenotypically normal. B) The SMAD2 rearrangement as
seen in our MPseq analysis pipeline. This junction plot displays four
breakpoints representing the inversion event (colored dots indicate
orientation of mapped reads, with blue corresponding to forward strand
and red the reverse strand). Notice the presence of the blue-blue and
red-red dot associations predicted to be seen in a duplication with
inverted orientation. Only discordant mate-pair reads with end positions
mapping >15 kb are shown. C) Schematic representation of
the SMAD2 duplication seen in proband compared to the reference
allele. Concordant and discordant mate-pair fragments and their
corresponding orientations are shown in colored text.
Figure 3. SMAD2 RNA expression analysis. Total mRNA was
isolated from cultured proband amniocytes, and RNA extracted from the
amniocytes of a normal male with normal SMAD2 copy number and no
phenotypic evidence of dextrocardia was used as a control. Real-time
RT-PCR analysis targeted the SMAD2 exon junctions 2/3, 4/5, 6/7,
and 9/10 indicated underneath the black horizontal bars in both the
proband (grey bars and Pt label) and the control (black bars and Ctrl
label). NTC: no template control. The expression of SMAD2 is
significantly reduced in proband compared to the normal control.