2.8 Real-time polymerase chain reaction (PCR)
Total RNA (1 μg) extracted from atrial tissue using the RNeasy Mini Kit
(Qiagen, Valencia, CA, USA) was reverse transcribed into cDNA using the
High-Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific,
Inc., Waltham, MA, USA), as previously described (Aonuma et al., 2020;
Feng et al., 2020; Xu et al., 2012; Xu et al., 2010). RT-PCR was
performed on the ABI Prism 7500 FAST sequence detection system (Applied
Biosystems, Foster City, CA, United States) using the PrimeTime Gene
Expression Master Mix (Integrated DNA Technologies). The following
primers were used: Nppb (Mm.PT.588584045.g), Col1a1 (Mm.PT.587562513),
Tgfb1 (Mm.PT.5811254750), Ryr2 (Mm.PT.58.45974879), Camkiia
(Mm.PT.58.30362101), and Cacna1c (Mm.PT.58.16361922). Gene expression
was normalized to that of the housekeeping gene, 18S rRNA (4319413E;
Thermo Fisher Scientific).