Processing human frontal cortex brain tissue for population-scale Oxford Nanopore long-read DNA sequencing SOP v1

2021 ◽  
Author(s):  
Kimberley J J Billingsley ◽  
Ramita Dewan ◽  
Laksh Malik ◽  
Pilar Alvarez Jerez ◽  
Stith Kiley ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Dna Sequencing ◽  
Frontal Cortex ◽  
Brain Tissue ◽  
Sequencing Data ◽  
Oxford Nanopore ◽  
Human Frontal Cortex ◽  
Long Read ◽  
Population Scale

Processing human frontal cortex brain tissue for population-scale Oxford Nanopore long-read DNA sequencing SOP At the NIH's Center for Alzheimer's and Related Dementias (CARD) https://card.nih.gov/research-programs/long-read-sequencing we will generate long-read sequencing data from roughly 4000 patients with Alzheimer's disease, frontotemporal dementia, Lewy body dementia, and healthy subjects. With this research, we will build a public resource consisting of long-read genome sequencing data from a large number of confirmed people with Alzheimer's disease and related dementias and healthy individuals. To generate this large-scale nanopore sequencing data we have developed a protocol for processing and long-read sequencing human frontal cortex brain tissue, targeting an N50 of ~30kb and ~30X coverage. †Correspondence to: Kimberley Billingsley [email protected] and Cornelis Blauwendraat [email protected] Acknowledgements: We would like to thank the Nanopore team (Androo Markham &Hannah Lucio), Circulomics Inc team (Jeffrey Burke, Michelle Kim, Duncan Kilburn & Kelvin Liu) and the whole CARD long-read team listed below => UCSC: Benedict Paten, Mikhail Kolmogorov, Miten Jain, Kishwar Shafin, Trevor Pesout; NHGRI: Adam Phillippy, Arang Rhie; Baylor: Fritz Sedlazeck; JHU: Winston Timp; NINDS: Sonja Scholz; NIA: Cornelis Blauwendraat, Kimberley Billingsley, Frank Grenn, Pilar Alvarez Jerez, Bryan Traynor, Shannon Ballard, Caroline Pantazis; CZI: Paolo Carnevali.

scholarly journals Systematic analysis of dark and camouflaged genes: disease-relevant genes hiding in plain sight

2019 ◽  
Author(s):  
Mark T. W. Ebbert ◽  
Tanner D. Jensen ◽  
Karen Jansen-West ◽  
Jonathon P. Sens ◽  
Joseph S. Reddy ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Sequencing Data ◽  
Systematic Analysis ◽  
Protein Coding ◽  
Short Read ◽  
Sequencing Project ◽  
Short Read Sequencing ◽  
Sequencing Technologies ◽  
Long Read

AbstractBackgroundThe human genome contains ‘dark’ gene regions that cannot be adequately assembled or aligned using standard short-read sequencing technologies, preventing researchers from identifying mutations within these gene regions that may be relevant to human disease. Here, we identify regions that are ‘dark by depth’ (few mappable reads) and others that are ‘camouflaged’ (ambiguous alignment), and we assess how well long-read technologies resolve these regions. We further present an algorithm to resolve most camouflaged regions (including in short-read data) and apply it to the Alzheimer’s Disease Sequencing Project (ADSP; 13142 samples), as a proof of principle.ResultsBased on standard whole-genome lllumina sequencing data, we identified 37873 dark regions in 5857 gene bodies (3635 protein-coding) from pathways important to human health, development, and reproduction. Of the 5857 gene bodies, 494 (8.4%) were 100% dark (142 protein-coding) and 2046 (34.9%) were ≥5% dark (628 protein-coding). Exactly 2757 dark regions were in protein-coding exons (CDS) across 744 genes. Long-read sequencing technologies from 10x Genomics, PacBio, and Oxford Nanopore Technologies reduced dark CDS regions to approximately 45.1%, 33.3%, and 18.2% respectively. Applying our algorithm to the ADSP, we rescued 4622 exonic variants from 501 camouflaged genes, including a rare, ten-nucleotide frameshift deletion in CR1, a top Alzheimer’s disease gene, found in only five ADSP cases and zero controls.ConclusionsWhile we could not formally assess the CR1 frameshift mutation in Alzheimer’s disease (insufficient sample-size), we believe it merits investigating in a larger cohort. There remain thousands of potentially important genomic regions overlooked by short-read sequencing that are largely resolved by long-read technologies.

scholarly journals A multi-omic atlas of the human frontal cortex for aging and Alzheimer’s disease research

2018 ◽  
Vol 5 (1) ◽  
Author(s):  
Philip L. De Jager ◽  
Yiyi Ma ◽  
Cristin McCabe ◽  
Jishu Xu ◽  
Badri N. Vardarajan ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Frontal Cortex ◽  
Disease Research ◽  
Human Frontal Cortex

scholarly journals LRP3, an apolipoprotein receptor that links reelin signalling and APP expression, is affected in Alzheimer's disease

2021 ◽  
Author(s):  
Inmaculada Cuchillo-Ibañez ◽  
Matthew P Lennol ◽  
Sergio Escamilla ◽  
Trinidad Mata-Balaguer ◽  
Inmaculada López-Font ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Frontal Cortex ◽  
Ldl Receptor ◽  
Full Length ◽  
Adult Brain ◽  
Western Blots ◽  
Protein Levels ◽  
Human Frontal Cortex ◽  
Receptor Family

Abstract Background. Members of the low-density lipoprotein (LDL) receptor family are involved in endocytosis and in transducing signals, but also in APP (amyloid precursor protein) processing and β-amyloid secretion. ApoER2/LRP8 is a member of this family with key roles in synaptic plasticity in the adult brain. ApoER2 is cleaved after the binding of its ligand, the reelin protein, generating an intracellular domain (ApoER2-ICD) that modulates reelin gene transcription itself. In this work, we have analysed whether ApoER2-ICD is able to regulate the expression of other members of the LDL receptor family. We focused on LRP3, the most unknown member of the LDL receptor family, whose precise physiological role and potential participation in pathological processes such as Alzheimer’s disease (AD) are still unknown.Methods. The effects of full-length ApoER2 and ApoER2-ICD overexpression on protein levels, in presence of recombinant reelin or Ab42 peptide, were evaluated by a microarray, qRT-PCRs and western blots. The expression of LRP3 was analysed in human frontal cortex extracts from AD and non-demented subjects by qRT-PCRs and western blot; and LRP3 interaction with other proteins was assessed by immunoprecipitation. In CHO cells overexpressing LRP3, protein levels of full-length APP and fragments were evaluated by western blots. Results. We have identified that ApoER2 overexpression increases LRP3 expression. Stimulation of ApoER2 signaling by reelin increased LRP3 levels, and the same occurred following ApoER2-ICD overexpression. In human frontal cortex extracts we demonstrate that LRP3 interacts with apolipoprotein E and APP. In extracts from AD subjects, the levels of LRP3 mRNA and protein were lower than those in control subjects. Interestingly, LRP3 transfection in CHO-PS70 cells induced a decrease of full-length APP levels and APP-CTF, and in the supernatant, levels of soluble APP fragments from the amyloidogenic (sAPPa) or non-amyloidogenic (sAPPβ) pathway, as well as Aβ peptides, were drastically reduced respect to mock-transfected cells.Limitations. There is a scarce knowledge of LRP3 physiological function as a neuronal receptor.Conclusion. We describe that LRP3 expression is regulated via ApoER2/reelin signaling, and its levels are affected in AD; similarly to other LDL receptors, LRP3 is involved in APP expression.

scholarly journals Genome-Wide DNA Methylation Differences Between Late-Onset Alzheimer's Disease and Cognitively Normal Controls in Human Frontal Cortex

2012 ◽  
Vol 29 (3) ◽  
pp. 571-588 ◽  
Author(s):  
Kelly M. Bakulski ◽  
Dana C. Dolinoy ◽  
Maureen A. Sartor ◽  
Henry L. Paulson ◽  
John R. Konen ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Dna Methylation ◽  
Frontal Cortex ◽  
Late Onset ◽  
Cognitively Normal ◽  
Genome Wide ◽  
Human Frontal Cortex ◽  
Normal Controls
Sign in / Sign up

Export Citation Format

Share Document