Engineering Material Properties of Transcription Factor Condensates to Control Gene Expression in HEK-293T Cells
Engineering Material Properties of Transcription Factor Condensates to Control Gene Expression in HEK-293T Cells
600,000 HEK-293T cells/well were seeded onto 6-well plates (Corning, catalog no.CORN3516) and 24 h later transfected with 600 µL transfection mix containing 3.750 µg total DNA and 12.375 µL PEI per well. Blue-light illumination (460 nm, 5 µmol m-2 s-1) was started 8 h after transfection for 24 h. Controls were kept in darkness. Cells were harvested 24 h after treatment using 1 mL TriFast™ (VWR, cat. no. 30-2010) per well. Total RNA was extracted according to the manufacturer's protocol. RNA was further cleaned up using an RNeasy Mini Kit (Qiagen, cat. no. 74104) according to the manufacturer's protocol, including an on-column DNase I digest using 50 µL reactions of DNase I (New England Biolabs, cat. no. M0303). RNA integrity was verified at BGI Tech Solutions (Hong Kong) Co. using a Bioanalyzer (Agilent 2100 Bioanalyzer). Strand-Specific mRNA Library Preparation (DNBSEQ). BGI Protocol.