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Exome sequences account for only 2% of the genome and may overlook mutations causing disease. To obtain a more complete view, whole genome sequencing (WGS) was analyzed in a large consanguineous family in which members displayed autosomal recessively inherited cerebellar ataxia manifesting before 2 years of age.WGS from blood-derived genomic DNA was used for homozygosity mapping and a rare variant search. RNA from isolated blood leukocytes was used for quantitative polymerase chain reaction (PCR), RNA sequencing, and comparison of the transcriptomes of affected and unaffected family members.WGS revealed a point mutation in noncoding RNA RNU12 that was associated with early onset cerebellar ataxia. The U12-dependent minor spliceosome edits 879 known transcripts. Reverse transcriptase PCR demonstrated minor intron retention in all of 9 randomly selected RNAs from this group, and RNAseq showed splicing disruption specific to all U12-type introns detected in blood monocytes from affected individuals. Moreover, 144 minor intron-containing RNAs were differentially expressed, including transcripts for 3 genes previously associated with cerebellar neurodegeneration.Interference with particular spliceosome components, including small nuclear RNAs, cause reproducible uniquely distributed phenotypic and transcript-specific effects, making this an important category of disease-associated mutation. Our approach to differential expression analysis of minor intron-containing genes is applicable to other diseases involving altered transcriptome processing. ANN NEUROL 2017;81:68-78.

Objective: Information that is present in our DNA is transferred to RNA, which is in turn transcribed into proteins. This is a central principle of molecular biology. Over the years, there have been many attempts to document the thousands of places where transcribed RNA does not match the DNA template it was created from. This behavior is called RNA Editing, and is generally thought to be mainly an A->I change, that is, adenosine to inosine (read as guanine by DNA sequencers), with occasional cytosine to uracil changes as well. Cancer, with its volatility due to DNA base changes, gets more complex with the occurrences of RNA editing. To ensure a fair ground to study RNA editing, we have considered exome and RNASeq library pairs from the cancer-affected ovarian, lymph node and peritoneum tissues in three individual. This data set will help identify RNA editing locations in these tissues, and the role it plays in metastasis. Methods: The exome libraries were prepared using Illumina Paired End Sample Preparation kit and Agilent SureSelect Target Enrichment System kit. RNASeq libraries were prepared using Illumina paired-end protocol. Both the libraries were sequenced using the Illumina NextGen Sequencing platform. Exome libraries were aligned using bowtie2 and RNASeq using tophat2. SNPs were called using samtools. The RNAEditing database from Ensemble was used to filter out known editing sites. With the remaining editing locations, a blat was run against the surrounding sequence to check for similarity to other parts of the genome, to rule out editing sites being called as a result of incorrect alignment scoring. Results: From our preliminary analysis, we see that a majority of the RNA Editing positions represent A->I changes, with a few C->U changes as well. Some of these positional changes are seen across all tissues in an individual. There are few RNA edited sites that are mainly seen only in the metastatic locations, and not seen in the primary. Conclusions: By analyzing the initial results, we can identify the mutations and RNA edited sites that originate from the primary tumor tissue, and those that are acquired in the metastatic tissues.

The date palm is one of the oldest cultivated trees and is critical to the development of arid land. The date palm is a dioecious monocot with separate male and female trees, however only the female trees produce the fruit that is sought after in farming. This presents a challenge in crop development, as it is impossible to distinguish trees until they flower approximately 5–8 years after planting. We have developed two Polymerase Chain Reaction (PCR) based assays capable of sex differentiation in multiple date palm cultivars. The primers are designed across gender specific polymorphisms and demonstrated greater than 90% accuracy in distinguishing date palm gender across multiple varieties. These assays should be helpful in rapidly distinguishing date palm gender from the earliest stages that DNA can safely be collected. As the sex-linked region is fine mapped these assays will be refined to take into account this information. For now, our assays provide a vast savings in time and effort over exi...