Genotyping Protocol
Venipuncture was performed and DNA was extracted using the Illustra
Nucleon BACC3 Genomic DNA Extraction kit (GE Healthcare, Chicago, IL,
USA). Samples were genotyped for the AF-associated variants using TaqMan
(Applied Biosystems, Foster City, CA), a plate-based, 1-step reaction
that identifies the SNP allele during polymerase chain reaction
amplification. An AF genetic risk score was calculated using the
literature risk estimates as weights for the minor allele of each of the
12 SNPs as previously reported4. All 12 SNPs were
genotyped in all subjects without genotype imputation. In the atrial
appendage cohort subgroup, we classified patients based on genetic risk
score as well as the status of the RS2200733 SNV (a component of the
polygenic risk score). This SNP was chosen as a stratifier for
morphologic evaluation analysis given the known role of PITx2 in
Left-Right signaling. Patients were categorized as SNV carriers if they
were heterozygous (CT) or homozygous (TT) for the RS2200733 variant.
Patients with the CC genotype were classified as non-carriers.
Investigators were fully blinded to the genotype status of each subject
until data analysis was complete.
Modeling of genetic risk
score
The polygenic risk score utilized here includes 12 SNPs (Table 1), and
is the most commonly studied such score as originally published by Tada
et al and subsequently validated3, 5-9. The linkage
disequilibrium between these SNPs is captured in the implementation of
the score components4. The AF- genetic risk score
(GRS) for each individual in the current study was calculated: for each
SNP the natural log transformed risk estimate for the minor allele was
multiplied by the number of minor alleles carried by that individual;
these 12 products were then summed. The risk estimate for the minor
allele at each SNP was obtained from the largest available data set.