Effect of siRNA depletion of SPRTN gene on gene expression in HepG2 cells
A biallelic mutation in the SPRTN gene is currently the only known single-gene mutation implicated in early-onset HCC, contributing to the development of liver cancer. The precise mechanism by which SPRTN mutations lead to HCC is not fully understood. The SPRTN protein is involved in repair of DNA damage caused by DNA-protein crosslinks (DPC). Spartan was silenced in HepG2 cells with siRNA that targets mRNA of Spartan using Lipofectamine RNAiMAX (Thermo Fisher).Total RNA was extracted with QIAzol (Qiagen) according to the manufacturer's protocol and transcribed into cDNA with High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems) according to the manufacturer's protocol to test level of silencing with RT-qPCR .  The quality of RNA was tested with agarose electrophoresis and concentration was determined using Nanodrop. Samples were send to company providing RNA sequencing. Company did RNA quality check, sequencing and analysis of the results. Overall design: siRNA knockdown of SPRTN gene (siRNA-SPRTN-1, 5'-GUC AGG AAG UUC UGG UUA A-UA-3')  in HepG2 cell line