Computational protocol: Intra-tumour molecular heterogeneity of clear cell renal cell carcinoma reveals the diversity of the response to targeted therapies using patient-derived xenograft models

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Protocol publication

[…] Snap frozen PDX tumour samples were used to extract RNA for transcriptome sequencing (RNA-seq). The purity and integrity were checked with an Agilent Bioanalyzer prior to RNA sequencing. Only RNA samples with RIN >7, 28S/18S >1 and mouse content <10% were used for library construction and sequencing. The sequencing was performed at PE125 on an Illumina HiSeq2500 platform (KI2367) and PE100 on an Illumina HiSeq2500 platform (KI2368) by certified Illumina service providers. The RNAseq raw data were first cleaned by removing reads that were preferentially mapped to a mouse genome (UCSC MM9). Transcript expression was estimated by MMSEQ [] and represented by log2(FPKM). SNP and INDELs were detected by STAR [] mapping software and GATK [] variant discovery toolkit, and gene fusions were detected by SOAP fuse [] and deFuse []. [...] All statistical analyses were performed using SPSS (IBM Corp. Released 2011. IBM SPSS Statistics for Windows, Version 20.0. Armonk, NY: IBM Corp.). Two sample t-tests were used to check the equality of means between two groups. One-way ANOVA was used to check the equality of means among 3 and more groups. Differences were considered statistically significant at P< 0.05. […]

Pipeline specifications

Software tools MMSEQ, STAR, GATK, deFuse, SPSS
Applications Miscellaneous, RNA-seq analysis
Organisms Homo sapiens, Homo sapiens/Mus musculus xenograft, Mus musculus
Diseases Carcinoma, Renal Cell, Neoplasms