Multiomics Analysis of Leukemic Cells

Leukemic cell mRNA and protein sequencing was performed as previously described (4 (link)). In brief, RNA was extracted using TRIzol reagent and quantitated by fluorimetry (Qubit), and integrity was assessed using a TapeStation 2200 (Agilent Technologies). RNA sequencing was performed using Truseq poly-A selected library construction (Illumina) and paired end 100-bp sequencing using Hiseq 2000 and 2500 sequencers. Proteins were extracted from pre-B cells and digested in solution by sequential addition of Lys-C and trypsin proteases. The resulting peptides were individually desalted and labeled with 6-plex Tandem Mass Tag reagents (Thermo Scientific) for sequencing by tandem mass tag-based mass spectrometry.

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Churchman M.L., Evans K., Richmond J., Robbins A., Jones L., Shapiro I.M., Pachter J.A., Weaver D.T., Houghton P.J., Smith M.A., Lock R.B, & Mullighan C.G. (2016). Synergism of FAK and tyrosine kinase inhibition in Ph+ B-ALL. JCI Insight, 1(4), e86082.

Publication 2016

TapeStation 2200 6-plex Tandem Mass Tag reagents

Cell Fluorimetry Library Mrna Peptides Poly a Pre b cells Proteases Proteins Tandem mass spectrometry Trizol Trypsin

Corresponding Organization : St. Jude Children's Research Hospital

Other organizations : UNSW Sydney, Cancer Institute of New South Wales, Verastem (United States), The University of Texas Health Science Center at San Antonio, National Cancer Institute

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Variable analysis

independent variables

RNA extraction method (TRIzol reagent)
RNA quantitation method (Qubit fluorimetry)
RNA integrity assessment method (TapeStation 2200, Agilent Technologies)
RNA sequencing method (Truseq poly-A selected library construction, Illumina, paired-end 100-bp sequencing using Hiseq 2000 and 2500 sequencers)
Protein extraction and digestion method (pre-B cells, Lys-C and trypsin proteases)
Peptide labeling method (6-plex Tandem Mass Tag reagents, Thermo Scientific)
Mass spectrometry method (tandem mass tag-based)

dependent variables

MRNA and protein sequencing data

control variables

Previously described experimental protocols (4)

controls

No specific positive or negative controls were mentioned in the provided information.

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