TY - JOUR T1 - Optimizing nucleic acid extraction and transcriptome evaluation from low-input, fixed clinical samples JF - bioRxiv DO - 10.1101/2020.06.02.122150 SP - 2020.06.02.122150 AU - Katie M. Campbell AU - Egmidio Medina AU - Ignacio Baselga Carretero AU - Yaroslav Teper AU - Rangasamy Elumalai AU - Raymond Miller AU - Kasturi Pal AU - Jessica Pitzer AU - Eugenio Daviso AU - Carsten Carstens AU - James Laugharn AU - Willy Hugo AU - Jia Ming Chen AU - Agustin Vega-Crespo AU - Sameeha Jilani AU - Ivan Perez Garcilazo AU - Ling Dong AU - Xinmin Li AU - Siwen Hu-Lieskovan AU - Antoni Ribas Y1 - 2020/01/01 UR - http://biorxiv.org/content/early/2020/06/03/2020.06.02.122150.abstract N2 - Tumor biopsies are commonly formalin-fixed and paraffin-embedded (FFPE) for long-term and efficient storage. However, FFPE preservation can greatly compromise the quality of samples, the extraction of nucleic acids, and feasibility of downstream studies, including RNA sequencing. These challenges are especially evident in the studies of clinical trial samples, where the sizes of biopsies often limit the amount of material available for study. Here, we evaluate two nucleic acid extraction kits (Covaris truXTRAC FFPE tNA Plus, QIAGEN miRNeasy FFPE) and three hybridized-capture-based RNA sequencing library preparations (Agilent SureSelect XT RNA Direct, Agilent SureSelect XT HS, and Illumina TruSeq RNA Exome) to evaluate the impact of these sample processing steps on transcriptome evaluation in a melanoma biopsy procured in a clinical setting and preserved by FFPE. While there exist many options for extraction and library preparation that are appropriate for RNA sequencing from FFPE samples, we observed that combinations of experimental approaches may have subtle impacts on downstream analysis, including gene expression quantification and fusion detection.Competing Interest StatementKMC is a shareholder in Geneoscopy LLC. KP, JP, and ED are employees of Covaris. JL is and employee and shareholder of Covaris. RM and CC are employees of Agilent Technologies. RE is an employee and shareholder of Agilent Technologies. AR has received honoraria from consulting with Amgen, Bristol-Myers Squibb, Chugai, Genentech, Merck, Novartis, Roche and Sanofi, is or has been a member of the scientific advisory board and holds stock in Advaxis, Apricity, Arcus Biosciences, Bioncotech Therapeutics, Compugen, CytomX, Five Prime, FLX-Bio, ImaginAb, Isoplexis, Kite-Gilead, Lutris Pharma, Merus, PACT Pharma, Rgenix and Tango Therapeutics, has received research funding from Agilent and from Bristol-Myers Squibb through Stand Up to Cancer (SU2C). ER -