SV-FLB_6_KCNH2_Acc_T618S
Homo sapiens
Protocols: Approximately 1x10^6 hiPSCs (cell line described here: https://hpscreg.eu/cell-line/LUMCi028-A) were rinsed once with PBS without Mg2+ and Ca2+ (PBS-/-) and dissociated to single cells with 1x TrypLE Select (ThermoFisher). The dissociation reagent was neutralized with culture medium (DMEM/F12, BSA 0.5%) and the pellet of cells was resuspended in PBS-/- and kept at -20°C until the genomic DNA extraction protocol was performed. Genomic DNA was extracted from hiPSCs using the High Pure PCR Template Preparation Kit (Roche) and treated with RNAse A (10 mg/ml, Invitrogen) according to the manufacturers' instructions. 500 ng of gDNA was fragmented by sonication, followed by Illumina library prep using the KAPA Hyper Kit and unique barcoding of each sample with IDT adapters (xGen UDI-UMI adapters). The samples were purified twice to remove any adapter dimers and fragments below 150bp. The adapter ligated samples were further PCR enriched using the p5 and p7 primers for 10x cycles using the KAPA HiFi mastermix.The Hyb Module box of the SureSelectQXT Target Enrichment for the Illumina Platform (Agilent) was used for hybridization and capturing of each sample separately. Briefly, 750 ng of the Illumina-prepped sample and 500ng of the biotinylated RNA capture probes were used in the reaction. A 15 cycle post capture PCR with the p5 and p7 primers was performed using the KAPA HiFi mastermix, with the resulting samples purified using 0.8x Ampure XP beads (Beckman Coulthier) and quantified using the Qubit 2.0 (ThermoFisher).
ENA-FIRST-PUBLIC
2022-09-22
ENA-LAST-UPDATE
2022-09-22
External Id
SAMEA110424408
INSDC center alias
LUMC
INSDC center name
Leiden University Medical Center
INSDC first public
2022-09-22T00:41:13Z
INSDC last update
2022-09-22T00:41:13Z
INSDC status
public
Submitter Id
E-MTAB-11971:SV-FLB_6_KCNH2_Acc_T618S
broker name
ArrayExpress
cell_line
LUMCi028-A
cell_type
induced pluripotent stem cell
common name
human
developmental stage
adult
disease
Familial long QT syndrome
genetic modification
NM_000238.4:c.1853C>G
genotype
genetically modified genotype
organism part
skin
progenitor cell type
fibroblast
sample name
E-MTAB-11971:SV-FLB_6_KCNH2_Acc_T618S