BioProject

Purpose: To reveal the cystogenesis regulatory mechanism by miRNA and miRNA targeted genes in ADPKD mouse models. Method (mRNA-seq): In order to construct cDNA libraries with the TruSeq RNA library kit, 1ug of total RNA was used. The protocol consisted of polyA-selected RNA extraction, RNA fragmentation, random hexamer primed reverse transcription and 100nt paired-end sequencing by Illumina HiSeq2000. The libraries were quantified using qPCR according to the qPCR Quantification Protocol Guide and qualified using an Agilent Technologies 2100 Bioanalyzer. The transcript counts in isoform level and gene level were calculated, and the relative transcript abundances were measured in FPKM (Fragments Per Kilobase of exon per Million fragments mapped) using Cufflinks. Method (common): Both miRNA-seq and RNA-seq were carried out using the kidney tissues from two mouse models at postnatal day 1, 3 and 7 in triplicate samples. Total RNA was isolated using TRIzol method according to manufacturer’s protocol (Invitrogen Life Technologies). Result: UIn RNA-seq analysis, total 3,040 transcripts in Pkd1f/f:HoxB7-cre mice and 2,470 transcripts in Pkd2f/f:HoxB7-cre mice were differentially expressed at indicated time points (FDR<0.05). In addition, we also identified 1,297 common transcripts in both mouse models. In miRNA analysis, total 243 of miRNAs were identified as differentially expressed genes while 130 miRNAs were common in both mouse models. Among common miRNAs, total 13 miRNAs including 11 upregulated and 2 downregulated miRNAs were selected based on comparison of expression patterns at each time point. Conclusion: Our study described the parallel integrated analysis of miRNA-seq and RNA-seq data and validated the expression, direct interaction and cystogenesis-related functions of key miRNAs and mRNA targets. Overall design: miRNA and mRNA profiles according to cystogenesis of postnatal day 1, 3 and 7 in two representative ADPKD mouse models were generated by deep sequencing, in triplicate, using Illumina HiSeq2000.