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Variant annotation is an important step in deciphering the functional impact of genomic variants and their association with diseases. In this study, we analyzed 80 pan-cancer cases that underwent comprehensive genomic testing and compared the auto-classified variant tiers among four globally-available software solutions for variant interpretation from Roche, SOPHiA GENETICS, QIAGEN, and Genoox. The results revealed striking differences in tier classifications, which are believed to be a result of several factors, including subjectivity in the AMP/ASCO/CAP guidelines, threshold settings for variant allele frequencies and population allele frequencies, as well as variation in disease ontologies. Although the software tools described here provide a time-saving and repeatable process for interpretation of genomic data, it is crucial to understand the nuances and various settings for these solutions, as they can strongly influence variant tier classifications and downstream management.

Moyamoya disease (MMD) is a rare disorder characterized by cerebrovascular occlusion and development of hemorrhage-prone collateral vessels. Approximately 10–12% of cases are familial, with a presumed low penetrance autosomal dominant pattern of inheritance. Diagnosis commonly occurs only after clinical presentation. The recent identification of the RNF213 founder mutation (p.R4810K) in the Asian population has made a significant contribution, but the etiology of this disease remains unclear. To further develop the variant landscape of MMD, we performed high-depth whole exome sequencing of 125 unrelated, predominantly nonfamilial, ethnically diverse MMD patients in parallel with 125 internally sequenced, matched controls using the same exome and analysis platform. Three subpopulations were established: Asian, Caucasian, and non-RNF213 founder mutation cases. We provided additional support for the previously observed RNF213 founder mutation (p.R4810K) in Asian cases (P = 6.01×10−5) that was enriched among East Asians compared to Southeast Asian and Pacific Islander cases (P = 9.52×10−4) and was absent in all Caucasian cases. The most enriched variant in Caucasian (P = 7.93×10−4) and non-RNF213 founder mutation (P = 1.51×10−3) cases was ZXDC (p.P562L), a gene involved in MHC Class II activation. Collapsing variant methodology ranked OBSCN, a gene involved in myofibrillogenesis, as most enriched in Caucasian (P = 1.07×10−4) and non-RNF213 founder mutation cases (P = 5.31×10−5). These findings further support the East Asian origins of the RNF213 (p.R4810K) variant and more fully describe the genetic landscape of multiethnic MMD, revealing novel, alternative candidate variants and genes that may be important in MMD etiology and diagnosis.

In order to identify novel somatic mutations associated with classic BCR/ABL1-negative myeloproliferative neoplasms, we performed high-coverage genome sequencing of DNA from peripheral blood granulocytes and cultured skin fibroblasts from a patient with MPL W515K-positive primary myelofibrosis. The primary myelofibrosis genome had a low somatic mutation rate, consistent with that observed in similar hematopoietic tumor genomes. Interfacing of whole-genome DNA sequence data with RNA expression data identified three somatic mutations of potential functional significance: i) a nonsense mutation in CARD6, implicated in modulation of NF-kappaB activation; ii) a 19-base pair deletion involving a potential regulatory region in the 5′-untranslated region of BRD2, implicated in transcriptional regulation and cell cycle control; and iii) a non-synonymous point mutation in KIAA0355, an uncharacterized protein. Additional mutations in three genes (CAP2, SOX30, and MFRP) were also evident, albeit with no support for expression at the RNA level. Re-sequencing of these six genes in 178 patients with polycythemia vera, essential thrombocythemia, and myelofibrosis did not identify recurrent somatic mutations in these genes. Finally, we describe methods for reducing false-positive variant calls in the analysis of hematologic malignancies with a low somatic mutation rate. This trial is registered with ClinicalTrials.gov (NCT01108159).

Retinitis pigmentosa is a phenotype with diverse genetic causes. Due to this genetic heterogeneity, genome-wide identification and analysis of protein-altering DNA variants by exome sequencing is a powerful tool for novel variant and disease gene discovery. In this study, exome sequencing analysis was used to search for potentially causal DNA variants in a two-generation pedigree with apparent dominant retinitis pigmentosa.Variant identification and analysis of three affected members (mother and two affected offspring) was performed via exome sequencing. Parental samples of the index case were used to establish inheritance. Follow-up testing of 94 additional retinitis pigmentosa pedigrees was performed via retrospective analysis or Sanger sequencing.A total of 136 high quality coding variants in 123 genes were identified which are consistent with autosomal dominant disease. Of these, one of the strongest genetic and functional candidates is a c.269A>G (p.Tyr90Cys) variant in ARL3. Follow-up testing established that this variant occurred de novo in the index case. No additional putative causal variants in ARL3 were identified in the follow-up cohort, suggesting that if ARL3 variants can cause adRP it is an extremely rare phenomenon.

DNA-fluorescence in situ hybridisation (DNA-FISH) allows visualisation of chromosome organisation and rearrangement. FISH probes are pools of short fluorescently labelled DNA fragments that are often produced from template plasmids that contain large genomic inserts. For effective sample penetration and target hybridisation it is critical that probe fragments are between 200 and 500bp. Production of these short probes requires significant optimisation and can be confounded access to expensive sonication equipment or inherent sequence features that influence enzymatic fragmentation or amplification. Here we demonstrate that effective FISH probes can be prepared without the need for optimisation of fragmentation using a cocktail of two the 4bp recognition sequence restriction enzymes CviQI and AluI. Method name: Rapid probe construction for Fluorescence in situ hybridization (FISH), Keywords: Fluorescence in situ hybridization, FISH, Cancer, Translocation, Probe

Three American English diphthongs involving onsets in the back of the vowel space were investigated by means of acoustic measurements, discriminant analysis, and perceptual experiments. These diphthongs, termed DOOR/WAR, TOY, and STAR, were observed in the speech of ten young women residing in southern Michigan. The diphthongs were elicited in a variety of /b/-, //-, and //-initial words (e.g. board, Boyd, and barred). Words with similar consonantal margins but different nuclei (e.g. bode, baud, and bod) were elicited, as well. The DOOR/WAR and TOY words were compared with bodeand baud(and similar words with initial // and //), while the STAR words were compared with baudand bod(and similar words with initial // and //). All three methods of investigation showed that the initial part of the DOOR/WAR and TOY diphthongs more closely resembles that of bodethan baud. On the other hand, the initial part of the STAR diphthong was shown to be intermediate between that of baudand that of bod. [ABSTRACT FROM AUTHOR]

1. Sustainable river management demands a strategic approach that renders even routine decisions highly complex if they are to be consistent, transparent and accountable. Under these circumstances, there is an increasing need for decision-support systems, although carefully structured guidelines and statements of best practice will continue to play a major supporting role.2.This support is likely to be particularly important in wide-ranging contexts, from routine decisions in determining land drainage consents, to complex decisions involving significant uncertainty. Sustainable Upland River Corridor Management Evaluation System (SURCoMES) has been developed as a prototype system to demonstrate the potential scope and problems for providing decision support in complex and uncertain scenarios.3.In its present form it focuses on the selection of management responses to problems of river bank erosion. It caters for uncertainty by providing guidance as to whether a particular decision is more or less sustainable than alternative options. It does not determine whether actions are absolutely sustainable or not.4.Because of the judgemental nature of the questions, underlying criteria and standards have been built into the system. These evaluations, are, in part, subjective, even when based on existing best practice and experience, so a Delphi technique approach has been used to capture professional opinion from the research sector and the Environment Agency which is responsible for river management in England and Wales.5.Decision support will undoubtedly have an important future in sustainable environmental management. The SURCoMES approach indicates potential applications, but the system requires further development and validation before it is implemented more widely. Copyright © 2002 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]