Posttranscriptional regulation RNA-binding proteins UV crosslinking eCLIP iCLIP.Ĭopyright © 2023 The Authors exclusive licensee Bio-protocol LLC. Revised iCLIP-1.5 protocol is more efficient and highly robust it provides higher coverage even for low-input samples. Allows quantitative analysis of genome-wide changes in protein-RNA interactions. iCLIP facilitates the identification of sequence motifs recognized by RBPs. iCLIP-seq provides precise positional and quantitative information on the RNA-binding sites of RBPs in living cells. Key features Identification of RNA-binding sites of RNA-binding proteins (RBPs) at nucleotide resolution. Herein, we describe a step-by-step procedure for our revised iCLIP-seq protocol, that we designate as iCLIP-1.5, and provide alternative approaches for certain difficult-to-CLIP proteins. Our revised protocol takes advantage of several features of the eCLIP procedure and also improves on certain steps of the original iCLIP method, including optimization of circularization of cDNA. We utilized a modified iCLIP method to identify RNA-binding sites for SP1 and several of the cleavage and polyadenylation complex subunits, including CFIm25, CPSF7, CPSF100, CPSF2, and Fip1. We have recently reported that transcription factor SP1 functions in the regulation of alternative cleavage and polyadenylation through direct RNA binding. Several variants of CLIP have been developed to improve its efficiency and simplify the protocol. The completion of the research infrastructure at the Charles University and the establishment of new laboratory for the evaluation of the antiparasitic targets (Drug Discovery), which will assist in bridging the gap between the basic research and the application sphere.Individual nucleotide resolution UV cross-linking and immunoprecipitation followed by high-throughput sequencing (iCLIP-seq) is a powerful technique that is used to identify RNA-binding proteins' (RBP) binding sites on target RNAs and to characterize the molecular basis of posttranscriptional regulatory pathways. the acquisition of the missing state-of-the-art instruments for the applicant and the partners.The main goal is to combine the research efforts of the parasitological teams of the Charles University (BIOCEV, Dept of Parasitology), Parasitology Institute ASCR and the University of Ostrava and to create a team with a critical concentration of research capacities with high quality infrastructure and experienced support project management. The results will be published in high quality scientific journals or patented. The knowledge gained will be used for the development of new therapeutic and diagnostic procedures. The proposed research project activities are part of the program: 'Virulence factors of parasites as targets for development of novel antiparasitic strategies' and logically structured into five subprograms. Integrated money clip for your bills and receipts. Holds up to 12 cards with a large card selection window for easy and quick access. The main objective of the proposed research program is to discover the virulence factors of important parasites of humans and livestock, and to determine their potential for the development of new therapeutic and diagnostic procedures. The I-CLIP Full Grain Leather is our signature wallet that started it all. limited participation with international projects and cooperation.lack of communication between the basic research and the application sphere.absence of some important modern instruments.Organizational-economic problems, which project solves, include typical research deficiencies in CR: Significant problems are disease vectors as Ticks and Phlebotominaes. Used chemotherapeutics are usually toxic and against number of them has originated resistance without having available alternative chemotherapeutics. Project solves the absence of significant means for the treatment important infections caused by parasites. The proposed research activities will have performed at the Faculty of Science, Charles University, which includes the laboratories of the European Centre of Excellence BIOCEV and partner laboratories of the Institute of Parasitology of Biology Centre of the Czech Academy of Sciences in České Budějovice and newly established, laboratories at the Faculty of Science, University of Ostrava.

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