269 Generation of Human iPSC lines derived from PLN R14del Cardiomyopathy Patients 10 In this study, whole blood was collected from individuals carrying the PLN-R14del mutation, and healthy family probands. The PLN-R14del carriers can be either non-symptomatic up to now (2A, 3A, and 10A), or display already severe clinical symptoms (6B, 7A, and 10B). The peripheral blood mononuclear cells (PBMCs) were reprogrammed by the CytoTune-iPS 2.1 Sendai Reprogramming Kit using the manufacturer's protocol in feeder-dependent culture conditions. The hiPSC clones displayed a normal human-induced pluripotent stem cell (hiPSC) morphology by light microscopy (Fig. 1A), and positive immunostaining for NANOG and OCT4 in all hiPSC colonies (Fig. 1B). To further confirm the expression of pluripotency genes was confirmed via RT-qPCR analysis for SOX2, MYC, NANOG, OCT4, and ZFP42 (Fig. 1C). Karyotyping of the hiPSC lines was carried out by a ddPCR iCS-Digital™ PSC 24 probes test (Stemgenomics), which will ensure to catch 90% of most frequent genomic defects at high resolution such as the sub-karyotypic 20q.11.21 amplification, which accounts for 20% of all hiPSC abnormalities detected worldwide.4 All the lines showed no abnormal copy number variants (CNVs) and the normal expression of chromosome X for either male (0-1) or female (1-2) copy number expression (Supplementary Fig. 1). Microbiological contaminations were investigated on a regular basis by the MycoAlertTM Mycoplasma Detection Kit and were all negative (data not shown). A directed cardiac differentiation assay revealed that mesodermal cell types were detectable in all the lines (Fig. 1D). Hereafter, the generated hiPSC-CMs showed the capacity for cardiac expansion (Fig. 1E) before cryopreservation.5 All investigations were performed around passages p16–p20. In summary, we describe the generation described the generation hiPSC lines generated of three symptomatic PLN-R14del individuals, three non-symptomatic PLN-R14del individuals, and two non-carrier family members. All eight of the generated hiPSC lines have passed almost all the tests or the test are undergoing (Table 2). After all the tests are performed, the hiPSC lines are a useful model to study PLN-R14del cardiomyopathy in vitro.
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