Bulk RNA sequencing of bone marrow cells from MDS-RS patients and healthy donors

Cells were lysed in RLT (Qiagen) + 40 mM dithiothreitol (Sigma-Aldrich) and RNA extraction was performed with RNeasy Micro Kit (Qiagen) with RNase-free DNase treatment according to the manufacturer’s protocol. RNA integrity numbers (RIN) were estimated using Agilent RNA 6000 Pico Kits (Agilent Technologies, CA, USA). A minimum RIN value of 6.5 was considered adequate.

Minibulk: Minibulk RNAseq was performed for assessment of cycloheximide treatment effects in CD34+ and GPA+ cell populations. The library preparation procedure was performed using the Xpress Genomics bulk RNA-seq kit v1, automated on a SP960 liquid handler (MGI Tech). In short, the library preparation procedure denatures RNA samples in presence of oligo-dT primer, which is followed by reverse transcription of RNA with a template-switching procedure and pre-amplification of full-length cDNA for 10 PCR cycles. cDNA was subsequently tagmented using Tn5 (TDE1 Tagment DNA Enzyme; Illumina) and reactions quenched after 10 min at 55 °C by addition of 0.2 % SDS (Sigma-Aldrich).

Tagmented DNA was indexed using custom dual-unique Nextera index primers in a 12 cycle PCR reaction. Indexed libraries were cleaned up using SPRI beads in 22% PEG8000 buffer and eluted in 12 µL H2O. The dataset consists of 2 folders: - Bulk_Main - Minibulk_Cycloheximide The folder Bulk_Main contains 67 GNU zipped fastq files, 1 tsv file, and 1 txt file.

The folder Minibulk_Cycloheximide contains 2 GNU zipped fastq files, and 4 txt files. The documentation file File_list_BulkMinibulk.txt contains a full list of the files in the dataset. The total size of the dataset is approximately 360 GB.