GSM8183508: subject 59-CF medium_only time 2 h; Homo sapiens; RNA-Seq
For each RNA extraction, one well of a 12-well plate with fourteen 4 µl drops of Matrigel containing organoids was harvested using 1 ml of cold RNAprotect Tissue Reagent (Qiagen, Hilden, Germany). Total RNA was extracted using the RNeasy Plus Micro Kit (Qiagen) after homogenisation using a Beadbug homogeniser (Benchmark Scientific, Sayreville, NJ, U.S.A.). The quantity and purity of extracted RNA was verified using a Nanodrop (ThermoFisher, Waltham, MA, U.S.A.), and RNA integrity was evaluated with Agilent RNA 6000 Nano Kit on a Bioanalyser (both Agilent Technologies, Santa Clara, CA, U.S.). All RNA samples had a RNA Integrity Number (RIN) between 8 and 10. Gene transcription profiles were obtained by 3'-mRNA sequencing. Sequencing libraries were prepared using the Collibri 3' mRNA Library Prep Kit (ThermoFisher) and sequenced unidirectionally with a read length of 75 bases on a NextSeq 500 instrument using the NextSeq500/550 High Output v2.5 sequencing kit (both Illumina, Foster City, CA, U.S.A.). All libraries were constructed using one batch of the Collibri kit, and sequenced together in a single run.