scholarly journals Nanopore sequencing of full-length BRCA1 mRNA transcripts reveals co-occurrence of known exon skipping events

2017 ◽  
Author(s):  
Lucy C de Jong ◽  
Simone Cree ◽  
Vanessa Lattimore ◽  
George AR Wiggins ◽  
Amanda B. Spurdle ◽  
...  
Keyword(s):  
Exon Skipping ◽  
Full Length ◽  
Future Research ◽  
Molecular Evidence ◽  
Nanopore Sequencing ◽  
Brca1 Mrna ◽  
Dna Sequence Variants ◽  
Transcript Structure ◽  
Single Transcript ◽  
Long Range Pcr

AbstractLaboratory assays evaluating the effect of DNA sequence variants on BRCA1 mRNA splicing may contribute to classification by providing molecular evidence. However, our knowledge of normal and aberrant BRCA1 splicing events to date has been limited to data derived from assays targeting partial transcript sequences. For the first time, we resolve the exon structure of whole BRCA1 transcripts using MinION nanopore sequencing of long-range PCR amplicons. Our study identified 32 BRCA1 isoforms, including 18 novel isoforms which comprised skipping of multiple contiguous and/or non-contiguous exons. Furthermore, we show that known BRCA1 exon skipping events, such as Δ(9,10) and Δ21, can co-occur in a single transcript, with some isoforms containing four or more alternative splice junctions. Our results highlight complexity in BRCA1 transcript structure that has not previously been described. This finding has key implications for predicting translation frame of splicing transcripts, important for interpreting the clinical significance of spliceogenic variants. Future research is warranted to quantitatively assess full length BRCA1 transcript levels, and to assess the application of nanopore sequencing for routine evaluation of potential spliceogenic variants.

scholarly journals Nanopore sequencing of full-length BRCA1 mRNA transcripts reveals co-occurrence of known exon skipping events

2017 ◽  
Vol 19 (1) ◽  
Author(s):  
Lucy C. de Jong ◽  
◽  
Simone Cree ◽  
Vanessa Lattimore ◽  
George A. R. Wiggins ◽  
...  
Keyword(s):  
Exon Skipping ◽  
Full Length ◽  
Nanopore Sequencing ◽  
Mrna Transcripts ◽  
Brca1 Mrna

scholarly journals Re-evaluation of the distributions of dystrophin and utrophin in sciatic nerve

1995 ◽  
Vol 312 (1) ◽  
pp. 309-314 ◽  
Author(s):  
E Fabbrizio ◽  
J Latouche ◽  
F Rivier ◽  
G Hugon ◽  
D Mornet
Keyword(s):  
Peripheral Nerves ◽  
Full Length ◽  
Molecular Evidence ◽  
Related Protein ◽  
Terminal Part ◽  
Nervous Structure ◽  
Chromosome 6Q24 ◽  
Dmd Gene ◽  
And Function ◽  
Role And Function

Differential expression of proteins belonging to the dystrophin family was analysed in peripheral nerves. In agreement with previous reports, no full-size dystrophin was detectable, only Dp116, one of the short dystrophin products of the Duchenne muscular dystrophy (DMD) gene. We used specific monoclonal antibodies to fully investigate the presence of utrophin, a dystrophin homologue encoded by a gene located on chromosome 6q24. Evidence is presented here of the presence of two potential isoforms of full-length utrophin in different nerve structures, which may differ by alternative splicing of the 3′-terminal part of the utrophin gene according to the specificities of the monoclonal antiobodies used. One full-length utrophin was co-localized with Dp116 in the sheath around each separate Schwann cell-axon unit, but the other utrophin isoform was found to be perineurium-specific. We also highlighted a potential 80 kDa utrophin-related protein. The utrophin distribution in peripheral nerves was re-evaluated and utrophin isoforms were detected at the protein level. This preliminary indication will require more concrete molecular evidence to confirm the presence of these two utrophin isoforms as well as the potential 80 kDa utrophin isoform, but the results strongly suggest that each isoform must have a specialized role and function within each specific nervous structure.

scholarly journals Fundamental questions about genes, inactivity, and chronic diseases

2007 ◽  
Vol 28 (2) ◽  
pp. 146-157 ◽  
Author(s):  
Frank W. Booth ◽  
Simon J. Lees
Keyword(s):  
Physical Activity ◽  
Chronic Diseases ◽  
Physical Inactivity ◽  
Gene Interaction ◽  
Future Research ◽  
Molecular Evidence ◽  
Metabolic Dysfunction ◽  
Physically Active ◽  
Early Disease Detection ◽  
Artery Disease

Currently our society is faced with the challenge of understanding the biological basis for the epidemics of obesity and many chronic diseases, including Type 2 diabetes. Physical inactivity increases the relative risk of coronary artery disease by 45%, stroke by 60%, hypertension by 30%, and osteoporosis by 59%. Moreover, physical inactivity is cited as an actual cause of chronic disease by the US Centers of Disease Control. Physical activity was obligatory for survival for the Homo genus for hundreds of thousands of years. This review will present evidence that suggests that metabolic pathways selected during the evolution of the human genome are inevitably linked to physical activity. Furthermore, as with many other environmental interactions, cycles of physical activity and inactivity interact with genes resulting in a functional outcome appropriate for the environment. However, as humans are less physically active, there is a maladaptive response that leads to metabolic dysfunction and many chronic diseases. How and why these interactions occur are fundamental questions in biology. Finally, a perspective to future research in physical inactivity-gene interaction is presented. This information is necessary to provide the molecular evidence required to further promote the primary prevention of chronic diseases through physical activity, identify those molecules that will allow early disease detection, and provide society with the molecular information needed to counter the current strategy of adding physical inactivity into our lives.

P146 Nanopore sequencing as a maturing platform for full-length HLA genotyping

2017 ◽  
Vol 78 ◽  
pp. 163
Author(s):  
Vineeth Surendranath ◽  
Gerhard Schoefl ◽  
Kathrin Lang ◽  
Hanneke W. van Deutekom ◽  
Erik Rozemuller ◽  
...  
Keyword(s):  
Full Length ◽  
Nanopore Sequencing ◽  
Hla Genotyping

Database for chicken full-length cDNAs

2007 ◽  
Vol 28 (2) ◽  
pp. 141-145 ◽  
Author(s):  
Yong Wang ◽  
Zhenggang Wang ◽  
Juan Li ◽  
Yajun Wang ◽  
Frederick C. C. Leung
Keyword(s):  
Research Work ◽  
Full Length ◽  
Open Reading Frames ◽  
Future Research ◽  
Animal Science ◽  
Full Length Cdna ◽  
Chicken Genome Sequence ◽  
Gene Information ◽  
Expressed Sequence ◽  
Human And Mouse

The generation of full-length cDNA databases is essential for functional genomics studies as well as for correct annotation of species genomic sequences. Human and mouse full-length cDNA projects have provided the biomedical research community with a large amount of gene information. Recent completion of the chicken genome sequence draft now enables a similar full-length cDNA project to be initiated for this species. In this report, we introduce the development of a chicken full-length cDNA database, which will facilitate future research work in this biological system. In this project, chicken expressed sequence tags (ESTs) were aligned onto human and mouse full-length cDNAs (or open reading frames) on the basis of their similarity. More than 588,000 chicken ESTs were aligned to ∼170,000 full-length human and mouse templates obtained from the NEDO, RIKEN, and MGC databases. Many of these templates have known biological functions, and their orthologous chicken genes in the EMBL database are also provided in our database, which is available at http://bioinfo.hku.hk/chicken/ . We will continue to collect known chicken full-length cDNAs to update the database for public use. The cDNA alignment results presented herein and on our database will be useful for animal science and veterinary researchers wishing to clone and to confirm full-length chicken cDNAs of interest.

scholarly journals Exon-skipping Splice Variants of Excitatory Amino Acid Transporter-2 (EAAT2) Form Heteromeric Complexes with Full-length EAAT2

2010 ◽  
Vol 285 (41) ◽  
pp. 31313-31324 ◽  
Author(s):  
Florian M. Gebhardt ◽  
Ann D. Mitrovic ◽  
Daniel F. Gilbert ◽  
Robert J. Vandenberg ◽  
Joseph W. Lynch ◽  
...  
Keyword(s):  
Amino Acid ◽  
Excitatory Amino Acid ◽  
Splice Variants ◽  
Exon Skipping ◽  
Amino Acid Transporter ◽  
Full Length ◽  
Excitatory Amino Acid Transporter ◽  
Acid Transporter

scholarly journals Identification of high confidence human poly(A) RNA isoform scaffolds using nanopore sequencing.

RNA ◽  
2021 ◽  
pp. rna.078703.121
Author(s):  
Logan Mulroney ◽  
Madalee Wulf ◽  
Ira Schildkraut ◽  
George Tzertzinis ◽  
John Buswell ◽  
...  
Keyword(s):  
Ionic Current ◽  
Rna Degradation ◽  
Full Length ◽  
Correct Identification ◽  
Nanopore Sequencing ◽  
High Confidence ◽  
Protein Coding ◽  
Current Signatures

Nanopore sequencing devices read individual RNA strands directly. This facilitates identification of exon linkages and nucleotide modifications; however, using conventional methods the 5′ and 3′ ends of poly(A) RNA cannot be identified unambiguously. This is due in part to the architecture of the nanopore/enzyme-motor complex, and in part to RNA degradation in vivo and in vitro that can obscure transcription start and end sites. In this study, we aimed to identify individual full-length human RNA isoform scaffolds among ~4 million nanopore poly(A)-selected RNA reads. First, to identify RNA strands bearing 5′ m7G caps, we exchanged the biological cap for a modified cap attached to a 45-nucleotide oligomer. This oligomer adaptation method improved 5′ end sequencing and ensured correct identification of the 5′ m7G capped ends. Second, among these 5′-capped nanopore reads, we screened for ionic current signatures consistent with a 3′ polyadenylation site. Combining these two steps, we identified 294,107 individual high-confidence full-length RNA scaffolds, most of which (257,721) aligned to protein-coding genes. Of these, 4,876 scaffolds indicated unannotated isoforms that were often internal to longer, previously identified RNA isoforms. Orthogonal data confirmed the validity of these high-confidence RNA scaffolds.

scholarly journals Full Length Transcriptome Highlights the Coordination of Plastid Transcript Processing

2021 ◽  
Author(s):  
Marine Guilcher ◽  
Arnaud Liehrmann ◽  
Chloé Seyman ◽  
Thomas Blein ◽  
Guillem Rigaill ◽  
...  
Keyword(s):  
Gene Expression ◽  
Molecular Mechanisms ◽  
Full Length ◽  
Nanopore Sequencing ◽  
Rna Seq ◽  
Plastid Gene ◽  
Plastid Gene Expression ◽  
Short Read ◽  
Transcript Processing

Plastid gene expression involves many post-transcriptional maturation steps resulting in a complex transcriptome composed of multiple isoforms. Although short read RNA-seq has considerably improved our understanding of the molecular mechanisms controlling these processes, it is unable to sequence full-length transcripts. This information is however crucial when it comes to understand the interplay between the various steps of plastid gene expression. Here, the study of the Arabidopsis leaf plastid transcriptome using Nanopore sequencing showed that many splicing and editing events were not independent but co-occurring. For a given transcript, maturation events also appeared to be chronologically ordered with splicing happening after most sites are edited.

scholarly journals Direct RNA nanopore sequencing of full-length coronavirus genomes provides novel insights into structural variants and enables modification analysis

2018 ◽  
Author(s):  
Adrian Viehweger ◽  
Sebastian Krautwurst ◽  
Kevin Lamkiewicz ◽  
Ramakanth Madhugiri ◽  
John Ziebuhr ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Rna Virus ◽  
Consensus Sequence ◽  
Full Length ◽  
Viral Rna ◽  
Genome Replication ◽  
Nanopore Sequencing ◽  
Human Coronavirus ◽  
Viral Rnas ◽  
Virus Genomes

Sequence analyses of RNA virus genomes remain challenging due to the exceptional genetic plasticity of these viruses. Because of high mutation and recombination rates, genome replication by viral RNA-dependent RNA polymerases leads to populations of closely related viruses, so-called 'quasispecies'. Standard (short-read) sequencing technologies are ill-suited to reconstruct large numbers of full-length haplotypes of (i) RNA virus genomes and (ii) subgenome-length (sg) RNAs comprised of noncontiguous genome regions. Here, we used a full-length, direct RNA sequencing (DRS) approach based on nanopores to characterize viral RNAs produced in cells infected with a human coronavirus. Using DRS, we were able to map the longest (~26 kb) contiguous read to the viral reference genome. By combining Illumina and nanopore sequencing, we reconstructed a highly accurate consensus sequence of the human coronavirus (HCoV) 229E genome (27.3 kb). Furthermore, using long reads that did not require an assembly step, we were able to identify, in infected cells, diverse and novel HCoV-229E sg RNAs that remain to be characterized. Also, the DRS approach, which circumvents reverse transcription and amplification of RNA, allowed us to detect methylation sites in viral RNAs. Our work paves the way for haplotype-based analyses of viral quasispecies by demonstrating the feasibility of intra-sample haplotype separation. Even though several technical challenges remain to be addressed to exploit the potential of the nanopore technology fully, our work illustrates that direct RNA sequencing may significantly advance genomic studies of complex virus populations, including predictions on long-range interactions in individual full-length viral RNA haplotypes.

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