Project description:Intraepithelial lymphocytes (IELs) constitute the largest lymphocyte population in the body and exhibit direct cytotoxic effector functions. How IELs are generated and maintained as effector T cells, however, is poorly understood. While significant attention has been given to the role of commensal microbiota in the intestine, the predominance of dietary components in the small intestine suggests that dietary antigens may regulate IEL generation and homeostasis. Here, we show that conventional TCRab + CD4 + and CD8 + IEL populations are present in normal numbers in germ-free (GF) mice which lack microbiota.However, these IELs are severely depleted in antigen-free (AF) mice, GF mice fed an amino acid diet devoid of proteins. Moreover, the few remaining IELs in AF mice lack effector function. Notably, while TCRab + CD8ab + IELs in adult GF mice can persist for prolonged periods, they lose their effector function when fed an AF diet. IL-12 presumably produced by intestinal dendritic cells plays a critical role in the maintenance of TCRab + CD8ab + IELs and their effector functions. Importantly, mice lacking functional dietary antigen-induced TCRab + CD8ab + IELs exhibit a defect in early protection against a foodborne pathogen, Listeria monocytogenes. Collectively, these findings support that dietary antigen exposure drives the generation of innate-like cytotoxic IELs thatprovide rapid and local protection against foodborne-pathogens.
Project description:Amplicon-based targeted re-sequencing analysis was performed in the patient-derived gliobastoma cell culture samples. For this purpose, genomic DNA (gDNA) was isolated and DNA libraries were prepared using the TruSeq Custom Amplicon Low Input (Illumina, Inc.) technology. By this, a pool of 375 amplicons was generated for each single sample in order to enrich for the target genes ATRX1, EGFR, IDH1, NF1, PDGFRA, PIK3CG, PIK3R1, PTEN, RB1 and TP53. Sequencing was performed on the Illumina MiSeq® next generation sequencing system (Illumina Inc.) and its 2 x 250 bp paired-end v2 read chemistry. The resulting reads were quality controlled and mapped against the human reference genome (hg19). For all samples, sequence variations of the amplified regions of interest in comparison to the human reference sequence were identified and filtered based on reliability.
Project description:Purpose: The goals of this study are to introduce a new genome editing tool, which has the higher editing scope than the original genome editing tools. Methods: First, we transfected PE2 (the original prime editing tool, prime editor2), PE3 (the original prime editing tool, prime editor3) and HOPE (the new tool we developed in this study) vectors into human cells, respectively. Then, we harvested the genomic DNA form the transfected cells and amplified the specified amplicons. Finally, we used targeted amplicon sequencing approach to compare the editing efficiency and presion of the new tool with the original reported tools. Results: Our new genome editing tool improves the editing efficiency of prime editing without increasing the risk of undesired indels formation. Conclusions: We deleveped a new genome editing tool to increase the likelihood of successful gene engineering.
Project description:This study will include priority foodborne pathogens and investigate different methods, based on NGS data, of analysing and clustering these for the purpose of improved surveillance.