Retroelements are considered as one of the important sources of genomic variability in modern humans. It is known that transposition activity of retroelements in germline cells generates new insertions in various genomic loci and sometimes results in genetic diseases. Retroelements activity in somatic cells is restricted by different cellular mechanisms; however, there is an evidence for it in some tissue types. Somatic insertions can trigger tumorigenesis or participate in normal functioning such as generation of neurons` plasticity. In spite of the rapid development of high-throughput sequencing methods a confident detection of somatic insertions is still quite a challenging task. That, in part, is due to the absence of adequate bioinformatic tools for the analysis of sequencing data. Here, we propose an advanced computational pipeline for the identification of somatic insertions in datasets generated by selective amplification and high-throughput sequencing of genomic regions flanking insertions of AluYa5. Particular attention is paid for the identification of various artifacts arising in course of library preparation and the parameters for their filtration. Pipeline sensitivity is confirmed by in silico experiments with artificial datasets. Using the proposed pipeline we remove at least 80% of artifacts and preserve 75% of potentially somatic insertions. The approaches used in this work can be applied for the study of other mobile elements insertion variability.

A new quantitative method for minimal residual disease monitoring based on next generation dequencing and digital PCR principal, A new quantitative method for minimal residual disease monitoring based on next generation dequencing and digital PCR principal

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignant neoplasm of the lymphocyte precursors that suffered malignant transformation arresting the lymphoid cell differentiation. Clinical studies revealed monoor, more rarely, oligoclonal nature of the disease. A precise identification of malignant clone markers is both the crucial stage of early diagnostics and the essential prognostic factor for therapeutic treatment. Here we present an improved system for unbiased detection of lymphoblastic clones in bone marrow aspirates of T-ALL patients. The system based on multiplex PCR of rearranged T-cell receptor locus (TRB) and straightforward sequencing of the resulted PCR fragments. Testing of the system on genomic DNA from Jurkat cell line and four clinical bone marrow aspirates revealed a set of unique TRB rearrangements that precisely characterize each of tested samples. Therefore, the outcome of the system produces highly informative molecular genetic markers for further monitoring of minimal residual disease in T-ALL patients.

Detection of minimal residual disease (MRD) is a powerful prognostic tool in many hematological malignancies including ALL. Several groups of markers are widely used to monitor the concentration of a malignant clone including the detection of 'clonal B-cell (BCR) or T-cell (TCR) gene receptor rearrangements in ALL. Identification of a clonal rearrangement specific for the malignant clone, which usually constitutes from 20 to 90% in the initial bone marrow sample is a relatively straightforward task. Tracking this rearrangement after therapy is more tricky as the concentration of malignant cells in the sample can be very low. Recent progress in MRD detection based on quantitative real-time PCR (qPCR) and highthroughput sequen- cing (HTS) allows to detect malignant clones present at a concentration of one per 10 and even per 10 cells. Although MRD detection is claimed to be able to accurately predict the outcomes of leukemia therapy in different clinical settings, the value of this information is sometimes corrupted by false-positive and false-negative results of MRD measurement. One of the potential causes of false-positive results is discussed below.

We investigated an in vitro system for the analysis of template switching effect of RNA-dependent DNA-polymerase during reverse transcription reaction. An RT-PCR-based approach was realized. An RNA sample was used as a template. The RNA contains a region with a stable secondary structure flanked by two homologous sequences. The described system can find use for testing overall performance of new and existing forms of the reverse transcriptase.