Project description:We cultured MCF10a-Snail-ER cells and induced EMT initiation with tamoxifen. A matched sequencing of their PolyA RNA was performed, using Illumina and direct RNA Oxford Nanopore sequencing technologies. Both generated datasets supported the development of hybrid bioinformatics tools.
Project description:Noroviruses cause immense sporadic gastroenteritis outbreaks worldwide. Upcoming genotypes, which are divided based on VP1 sequence, further enhance this public thread regularly. Self-assembling properties of the human norovirus major capsid protein VP1 are crucial for using virus-like particles (VLPs) for vaccine development. However, there is no vaccine available yet. Here, VLPs from different variants produced in insect cells are characterized in detail using a set of biophysical and structural tools.
Project description:Background:During the winter of 2014-2015, a rarely reported norovirus (NoV) genotype GII.17 was found to have increased its frequency in norovirus outbreaks in East Asia, surpassing the GII.4 NoV infections. GII.17 genotype has been detected for over three decades in the world. The aim of this study is to examine the evolutionary dynamics of GII.17 over the last four decades. Methods:NoV GII.17 sequences with complete or nearly complete VP1 were downloaded from GenBank and the phylogenetic analyses were then conducted. Results:The maximum likelihood analysis showed that GII.17 genotype could be divided into four different clades (Clades A-D). The strains detected after 2012, which could be the cause of the outbreaks, were separated into Clades C-D with their mean amino acid distance being 4.5%. Bayesian Markov chain Monte Carlo analyses indicated that the rate of nucleotide substitution per sites was 1.68 × 10-3 nucleotide substitutions/site/year and the time of the most recent common ancestor was 1840. The P2 subdomain of GII.17 was highly variable with 44% (56/128) amino acids variations including two insertions at positions 295-296 and one deletion at position 385 (Clades C and D) and one insertion at position 375 (Clade D). Variations existed in Epitopes A, B and D corresponding to GII.4 and human histo-blood group antigens binding site I in P2 subdomain. Conclusion:The novel GII.17 strains that caused outbreaks in 2013-2015 may have two new variants. The evolvement of HBGAs binding site and epitopes in P2 subdomain might contribute to the novel GII.17 strains predominance in some regions.
Project description:Noroviruses are the principal cause of epidemic gastroenteritis worldwide. Multiple reports have concluded that the major capsid proteins of GII.4 strains, which cause 80% of norovirus infections worldwide, are evolving rapidly, resulting in new epidemic strains. Surrogate neutralization assays using sera from outbreaks and from immunized mice suggest that, as with influenza virus, antigenic variation maintains GII.4 persistence in the face of human population herd immunity. To test this hypothesis, mice were hyperimmunized with virus-like particles (VLPs) representing an early (GII.4-1987) and a contemporary (GII.4-2006) GII.4 strain. Anti-GII.4-1987 IgG monoclonal antibodies (MAbs) strongly reacted with GII.4 VLPs derived between only 1987 and 2002. Ligand binding blockade was more efficient with GII.4-1987 and GII.4-1997 VLPs than with GII.4-2002. Anti-GII.4-2006 IgG MAbs recognized either a broad panel of GII.4 VLPs (1987 to 2006) or a subset of contemporary (2004 to 2006) VLPs. Most 2006 antibodies did not recognize or only poorly recognized GII.4 VLPs of 2007 or 2008, documenting rapid antigenic evolution of GII.4 capsids. Generally, 2006 MAbs blocked homotypic VLP-ligand binding but were unable to block VLPs representing strains primarily circulating during or earlier than 2002. These analyses demonstrate that both subtle and significant evolutionary change has occurred within antibody epitopes between epidemic strains, providing direct evidence that the GII.4 noroviruses are undergoing antigenic variation, likely in response to herd immunity. As with influenza virus, HIV, and hepatitis C virus, norovirus antigenic variation will significantly influence the design of efficacious vaccines and immunotherapeutics against these important human pathogens.
Project description:Transposon insertion site sequencing (TIS) is a powerful method for associating genotype to phenotype. However, all TIS methods described to date use short nucleotide sequence reads which cannot uniquely determine the locations of transposon insertions within repeating genomic sequences where the repeat units are longer than the sequence read length. To overcome this limitation, we have developed a TIS method using Oxford Nanopore sequencing technology that generates and uses long nucleotide sequence reads; we have called this method LoRTIS (Long Read Transposon Insertion-site Sequencing). This experiment data contains sequence files generated using Nanopore and Illumina platforms. Biotin1308.fastq.gz and Biotin2508.fastq.gz are fastq files generated from nanopore technology. Rep1-Tn.fastq.gz and Rep1-Tn.fastq.gz are fastq files generated using Illumina platform. In this study, we have compared the efficiency of two methods in identification of transposon insertion sites.