Exome sequencing from nanogram amounts of starting DNA: comparing three approaches

Hybridization-based target enrichment protocols require relatively large starting amounts of genomic DNA, which is not always available. Here, we tested three approaches to pre-capture library preparation starting from 10 ng of genomic DNA: (i and ii) whole-genome amplification of DNA samples with REPLI-g (Qiagen) and GenomePlex (Sigma) kits followed by standard library preparation, and (iii) library construction with a low input oriented ThruPLEX kit (Rubicon Genomics). Exome capture with Agilent SureSelectXT2 Human AllExon v4+UTRs capture probes, and HiSeq2000 sequencing were performed for test libraries along with the control library prepared from 1 µg of starting DNA. Tested protocols were characterized in terms of mapping efficiency, enrichment ratio, coverage of the target region, and reliability of SNP genotyping. REPLI-g- and ThruPLEX-FD-based protocols seem to be adequate solutions for exome sequencing of low input samples

Student’s class

The article presents the results of the implementation in the educational processadditional classes on medical subjects.В статье представлены результаты внедрения в образовательный процесс школьников 9–11-х классов внеклассных занятий на медицинскую тематику

Whole exome sequencing links dental tumor to an autosomal-dominant mutation in ANO5 gene associated with gnathodiaphyseal dysplasia and muscle dystrophies

Tumors of the jaws may represent different human disorders and frequently associate with pathologic bone fractures. In this report, we analyzed two affected siblings from a family of Russian origin, with a history of dental tumors of the jaws, in correspondence to original clinical diagnosis of cementoma consistent with gigantiform cementoma (GC, OMIM: 137575). Whole exome sequencing revealed the heterozygous missense mutation c.1067G \u3e A (p.Cys356Tyr) in ANO5 gene in these patients. To date, autosomal-dominant mutations have been described in the ANO5 gene for gnathodiaphyseal dysplasia (GDD, OMIM: 166260), and multiple recessive mutations have been described in the gene for muscle dystrophies (OMIM: 613319, 611307); the same amino acid (Cys) at the position 356 is mutated in GDD. These genetic data and similar clinical phenotypes demonstrate that the GC and GDD likely represent the same type of bone pathology. Our data illustrate the significance of mutations in single amino-acid position for particular bone tissue pathology. Modifying role of genetic variations in another gene on the severity of the monogenic trait pathology is also suggested. Finally, we propose the model explaining the tissue-specific manifestation of clinically distant bone and muscle diseases linked to mutations in one gene

COMPARATIVE ANALYSIS OF HOSPITAL PHARMACOTHERAPY OF CHRONIC HEART FAILURE WITH REDUCED LEFT VENTRICULAR EJECTION FRACTIONS IN 2009-2010 AND 2014-2015

Aim. To compare features of pharmacotherapy of patients with chronic heart failure (CHF) with a reduced ejection fraction of the left ventricle (LV EF) who were admitted in a specialized department of the multidisciplinary hospital in Saratov before and after the publication of the Russian National Recommendations (4 revision) on the diagnosis and treatment of CHF (2012). Material and methods. A pharmacoepidemiological retrospective study was conducted. The object of the study was the medical records of inpatients (form 003/y) with the diagnosis "Heart failure" (ICD-I50), that consecutively admitted to the cardiology department of the multidisciplinary hospital in Saratov from April 28, 2009 to January 19, 2010 (n=52) and from February 19, 2014 to May 20, 2015 (n=95). Patients over 18 years of age with diagnosis of CHF (NYHA II-IV) and LV EF <45% were enrolled into the analysis. For each patient, an individual registration card was filled in which the patient's clinical and demographic characteristics, prescribed medications, their daily dose, the frequency of administration, the route of administration were indicated. Pharmacoepidemiological analysis was carried out for the drugs prescribed at the 1st day of hospitalization, at the 3rd-6th day (the time of stabilization of the patient's condition, which was evaluated by the reduction in dyspnea and increase in the tolerance to physical loads). The recommendations given by the physicians at discharge of the patients from the hospital were also considered. Results. In 2014-2015 years, compared to 2009-2010 years, the number of identified arrhythmias and severe forms of arterial hypertension significantly (82.1 vs 77%; р<0.05) increased. In 2014-2015 the frequency of the prescriptions of ACE inhibitors decreased (77.8 vs 86.5%; p<0.05). The frequency of the prescriptions of angiotensin II receptor blockers, antagonists of mineralocorticoid receptors (AMCR), diuretics, oral anticoagulants, clopidogrel increased (p<0.05). In the structure of combination therapy in 2014-2015, the frequency of the prescription of the ACE inhibitor+beta-blocker and ACE inhibitor+beta-adrenoblocker+AMCR combinations decreased significantly (18.9 vs 26.9%, p<0.05 and 22.1 vs 42.3%, p<0.05, respectively). At the same time prescription frequency of the ACE inhibitor+beta-blocker+AMCR+diuretic combination increased (25.2 vs 11.5%, p<0.05). Conclusion. Pharmacotherapy of CHF in hospital in 2014-2015 is consistent with the Russian National Recommendations (4 revision) and is significantly different from the CHF therapy in 2009-2010. Keywords: pharmacoepidemiology, chronic heart failure, treatment

The development of a method for determining the biological activity of erythropoietin preparations in vitro

The method for the assessment of biological activity of erythropoietin preparations in vitro on TF-1 sensitive cells culture has been developed. The special characteristics of measurements and calculations of the biological activity of erythropoietin with different levels of glycosylation in vivo and in vitro. Linear response ranges for cell lines to erythropoietin and its hyperglycosylated analogue differ and make (0.39-0.56 ng/ml) and (3.1-12.5 ng/ml), respectively. The results have shown the possibility of using cellular test for comparing and assaying the activity of erythropoietin analogues in standard international units of biological activity (IU) set for erythropoietin

Разработка метода определения биологической активности препаратов эритропоэтина in vitro

The method for the assessment of biological activity of erythropoietin preparations in vitro on TF-1 sensitive cells culture has been developed. The special characteristics of measurements and calculations of the biological activity of erythropoietin with different levels of glycosylation in vivo and in vitro. Linear response ranges for cell lines to erythropoietin and its hyperglycosylated analogue differ and make (0.39-0.56 ng/ml) and (3.1-12.5 ng/ml), respectively. The results have shown the possibility of using cellular test for comparing and assaying the activity of erythropoietin analogues in standard international units of biological activity (IU) set for erythropoietin.Разработан метод оценки биологической активности препаратов эритропоэтина in vitro на культуре чувствительных клеток TF-1. Исследованы особенности измерения и расчета биологической активности эритропоэтинов с различным уровнем гликозилирования в тестах in vivo и in vitro. Диапазоны линейного ответа клеточной линии на эритропоэтин и его гипергликозилированный аналог отличались и составляли (0,39-0,56 нг/мл) и (3,1-12,5 нг/мл) соответственно. Проведенный анализ полученных результатов показывает возможность применения клеточного теста для сопоставления и количественной оценки активности аналогов эритропоэтина в международных единицах стандарта биологической активности (МЕ), принятых для эритропоэтина