GSE125442 ( lncRNA+mRNA )

renal cell carcinoma ( renal )

Urinary exosomal long noncoding RNAs may be novel biomarkers for the diagnosis of clear cell renal cell carcinoma

Cancer : 10 / Control : 10

Illumina Bcl2FastQ software used for basecalling. Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence. The remaining reads were filtered against the rRNA database to remove possible ribosomal RNA contamination, and then mapped to the hg19 whole genome using Tophat2 v2.0.13 with parameters --read_mismatches 2 --read_gap_length 2 --read_edit_dist 2. Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.

Exosomes were isolated by ultracentrifugation from 50-ml urine samples from 10 patients with clear cell renal cell carcinoma and 10 matched healthy donors. Differentially expressed lncRNAs were analyzed by next-generation sequencing and further validated in kidney cancer cell lines, tissues and urinary exosomes. Then, we evaluated the sensitivity, specificity, clinical diagnostic value and stability of the selected lncRNAs.

Exosomes were isolated by ultracentrifugation from 50-ml urine samples from 10 patients with clear cell renal cell carcinoma and 10 matched healthy donors. Differentially expressed lncRNAs were analyzed by next-generation sequencing.

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A MiRNeasy Serum/Plasma Kit (QIAGEN, Valencia, CA, USA) was used to extract RNA from urinary exosome samples in accordance with the manufacturer’s protocol.

total RNA

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