September 27, 2013. New release of Shannon pipeline for detection of mRNA splicing mutations

Posted on September 29, 2013March 13, 2021 by Peter Rogan

Cytognomix has released a new version, V2.0, of the Shannon pipeline for mRNA splicing mutation analysis. It is distributed by CLC bio for use with their Genomics workbench or server. Click here to download the updated documentation.

This release (2.0):

is 12x faster (complete genome in 10-15 min) than version 1.0 and has fewer dependencies

is compatible and can be installed from the latest and previous versions of the CLC bio Workbench (6.5) and Server (5.0) software
now runs under Windows OS, (in addition to Linux and Mac iOS)
now handles insertions and deletions, (in addition to SNP snd DNPs),
performs gene set, ie. pathway, analysis on the genes found to contain mutations
has been updated to Ensembl v. 71 annotations
results can be optionally limited to Refseq genes,
and contains preset mutation filters to simplify analyses.

For laboratories that do not use CLC bio software, we have developed standalone version of the software which is available for custom integration into your existing NGS software analysis pipeline. It does not require any CLC bio products, but produces the same results as those products. Please contact Cytognomix if your are interested in this capability.

September 19. Abstract on metaphase epigenetics: platform presentation at American Society of Human Genetics meeting

Posted on September 20, 2013March 13, 2021 by Peter Rogan

Non-random, locus-specific differences in DNA accessibility are present in homologous metaphase chromosomes. W. A. Khan^1,3, P. K. Rogan^2,3,4, J. H. M. Knoll^1,3,4 1) Department of Pathology; 2) Departments of Biochemistry and Computer Science; 3) University of Western Ontario, London, Ontario, Canada; 4) Cytognomix, London, Ontario, Canada.
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Condensation differences between heterochromatin and euchromatin along the lengths of homologous, mitotic metaphase chromosomes are well known. This study describes differences in metaphase compaction between homologous euchromatic loci. We report molecular cytogenetic data showing local differences in condensation between homologs that are related to differences in accessibility (DA) of associated DNA probe targets. Reproducible DA was observed at ~10% of 450 distinct genomic regions mapped by single copy fluorescence in situ hybridization (scFISH). Fourteen short (1.5-5kb) sc and low copy (lc) FISH probes (from chromosomes 1, 5, 9, 11, 15, 16, 17, 22) targeting genic and non-genic regions with and without DA were developed and hybridized to cells from 10 individuals with cytogenetically-distinguishable homologs. Differences in hybridization were non-random for 6 genomic regions (RGS7, CACNAB1, HERC2, PMP22:IVS3, ADORA2B:IVS1, ACR) and were significantly-biased towards the same homolog (p< 0.01; n = 355 cells). The imprinted paternal chromosome 15 in a three-generation pedigree also showed non-random bias in DA. DNA probes within CCNB1, C9orf66, ADORA2B:Ex 1-IVS1, PMP22:IVS4-Ex 5, and a nongenic region within 1p36.3 did not show DA, while OPCML showed unbiased DA. A subset of probes was mapped onto chromosome topography by FISH-correlated atomic force microscopy (AFM). To quantify DA and pinpoint probe locations, we performed 3D-structured illumination super-resolution microscopy (3D-SIM). 3D anaglyph videos showed genomic regions with DA having nearly 5-fold larger differences in volumetric integrated probe intensities between homologs. Additional non-DA probes (NOMO1, NOMO3) hybridized to grooves in chromosome topography and exhibited a narrow range of probe depths (average: 0.08 μm) along axial and lateral axes of the 2 homologs. In contrast, probe for targets with DA (HERC2, PMP22:IVS3, ACR) significantly differed in probe depth (average: 0.77 μm) and volume (p < 0.05) between each homolog. Interestingly, genomic regions without DA are enriched in epigenetic marks (DHS, H3K27Ac, H3K4me1) of accessible interphase chromatin to a greater extent than regions with DA, suggesting these differences may be correlated with epigenetic marks established during the previous interphase. In summary, we present several lines of evidence that regional differences in condensation between homologs are programmed during metaphase chromosome compaction.

Click here for presentation details: session, location, time.

September 16, 2013. US patent to be awarded on breakthrough radiation biodosimetry technology

Posted on September 17, 2013March 29, 2014 by Peter Rogan

Cytognomix has received a Notice of Allowance from the US Patent and Trademark office for our patent application, “Centromere Detector and Method for Determining Radiation Exposure From Chromosome Abnormalities.” All claims that were applied for were allowed in the soon to be issued patent.

The application is available at this link: US Patent Application Ser. No 13/822,289

The application was originally filed November 4, 2011 as international patent application: PCT/US11/59257. This application was deemed patentable after examination, and was entered into the Patent Prosecution Highway program. Other applications remain pending in other jurisdictions.

September 6, 2013. Shannon pipeline highlighted at meeting on Clinical Applications of Next Generation Sequencing

Posted on September 6, 2013March 29, 2014 by Peter Rogan

Cytognomix’s Shannon pipeline has been highlighted in a report published describing the 2013 annual scientiﬁc meeting of the Human Genome Variation Society (HGVS) in Paris, France. The paper’s authors, Sian Ellard,George P. Patrinos, and William S. Oetting, write:

“Although NGS can identify numerous sequence variants, a major problem is predicting the functional consequence of mutations identiﬁed after sequencing. Much work has been done on determining the functionality of protein coding mutations within exon sequences, but signiﬁcant mutations that affect splicing or gene expression have not been comprehensively assessed. Peter Rogan, of the Schulich School of Medicine, Western University, London,Canada, has taken on this task and presented some of his results in his talk “Genome-wide prediction and validation of mRNA splicing mutations in cancer.” Using Shannon information theory, software packages were created for genome-scale analysis of variants within splice sites that alter binding site strength [Shirley et al., 2013] and to predict the transcript isoforms generated by splicing mutations [Mucaki et al., 2013]. Variants within the BRCA1 and BRCA2 genes were used to test their model and were grouped into four functional states: wild type, leaky mutations with residual activity, inactivating mutations, and cryptic splicing. Predicted effects of 50% of these calls were conﬁrmed using RNAseq analysis. Additional efforts underway include predicting functional mutations in promoter and intronic regions and the potential effects of mutations in regulatory splicing elements with information theory-based models. The goal is to use these software tools to perform a genome-scale mutation analysis to identify functional variants outside the coding regions affecting either splicing or expression levels.”

Citation: Ellard S, Patrinos GP, and Oetting WS, Clinical Applications of Next-Generation Sequencing: The 2013 Human Genome Variation Society Scientiﬁc Meeting, Human Mutation, DOI: 10.1002/humu.22400, 2013.