GSE118505
RNA-Seq mapping enabled quantitative analysis of gene expression differences between WT and rbm48 mutants in Zea mays (Maize).
Purpose: The goals of this study are to compare NGS-derived transcriptomes (RNA-Seq) derived from rbm48 mutant and WT maize endosperm tissue. Homozygous rbm48 mutants exhibit a seed defect and are seedling lethal. Maize Rbm48 is homologouse to a human protein (rbm48) that is  thought o be invoilved in the splicing of minor (U12) introns.  This study characterizes DEGs between rbm48 alleles umu1 and umu2 and their WT counterparts and serves to identify U12 introns exhibiting retention as evidenced by RNA-seq read density within U12 introns in rbm49 mutants relative to WT controls. Methods: Endosperm mRNA profiles from 16-18 days after pollination (DAP) normal and rbm48 kernels in the Zea mays W22 background were generated by deep RNA-Seq. Total RNA was prepared for 4 biological replicates of paired rbm48 and normal sibling pools.  4 total RNA samples of rbm48 umu1 and their WT controls, and 4 samples of rbm48 umu2 and their WT controls were obtained (16 total). Non strand specific Tru-Seq illumine libraries were constructed and sequenced on the Illumina HiSeq. The sequence reads that passed quality filters were aligned with GSNAP, read counts/gene were determined with the HTSeq-Count utility in the HTSeq package and DEG analysis performed with DESeq2.  Counts to U2 and U12 introns were also assessed and intron retention was assessed by measuring the Percent Splice Out (PSO) proportions for individual introns between mutant and WT samples. (PSO) calculations are based on the established methods for examining exon skip proportions (PSI). Conclusions: Differential gene expression analysis of rbm48 identifies large-scale changes. Considering counts in introns, the major U2-type introns are largely unaffected with only 3-5% of introns in expressed genes have a ?PSO >20%  By contrast, 65% and 53% of U12-type introns in rbm48-umu1 and rbm48-umu2, respectively, have a ?PSO >20% suggesting more than half of U12-type introns are retained in rbm48 mutants. These RNA-Seq data help demonstrate that rbm48 plays a role in splicing U12 introns. Overall design: Endosperm mRNA profiles from 16-18 days after pollination (DAP) normal and rbm48 kernels in the Zea mays W22 background were generated by deep RNA-Seq. Total RNA was prepared for 4 biological replicates of paired rbm48 and normal sibling pools (16 samples total). 2x100bp PE sequencing performed on an Illumina HiSeq2000.