Automated Solid-Phase Subcloning Based on Beads Brought into Proximity by Magnetic Force

In the fields of proteomics, metabolic engineering and synthetic biology there is a need for high-throughput and reliable cloning methods to facilitate construction of expression vectors and genetic pathways. Here, we describe a new approach for solid-phase cloning in which both the vector and the gene are immobilized to separate paramagnetic beads and brought into proximity by magnetic force. Ligation events were directly evaluated using fluorescent-based microscopy and flow cytometry. The highest ligation efficiencies were obtained when gene- and vector-coated beads were brought into close contact by application of a magnet during the ligation step. An automated procedure was developed using a laboratory workstation to transfer genes into various expression vectors and more than 95% correct clones were obtained in a number of various applications. The method presented here is suitable for efficient subcloning in an automated manner to rapidly generate a large number of gene constructs in various vectors intended for high throughput applications

ТРАНСПЛАНТАЦИЯ СЕРДЦА У ПАЦИЕНТКИ МОЛОДОГО ВОЗРАСТА С ДИЛАТАЦИОННОЙ КАРДИОМИОПАТИЕЙ И ВТОРИЧНЫМ АНТИФОСФОЛИПИДНЫМ СИНДРОМОМ

Patients with congestive heart failure have an increased incidence of thromboembolic events. The choice of me- dical management in patients with antiphospholipid antibodies and generalized thrombosis due to hypercoagula- bility is complex issue. We report heart transplant outcome in 15 years old patient with dilated cardiomyopathy and secondary anti-phospholipid syndrome. Пациенты с хронической сердечной недостаточностью имеют повышенный риск тромбоэмболических осложнений. Выбор врачебной тактики особенно затруднен при выявлении у таких больных антифосфо- липидных антител и развитии генерализованных тромбозов. В статье представлены результаты успеш- ной ортотопической трансплантации сердца у пациентки 15 лет с дилатационной кардиомиопатией и вторичным антифосфолипидным синдромом.

ТРАНСПЛАНТАЦИЯ СЕРДЦА У ПАЦИЕНТКИ С ПОСТТРАВМАТИЧЕСКОЙ АНЕВРИЗМОЙ ЛЕГОЧНОЙ АРТЕРИИ

Right heart catheterization (RHC) should be performed on all candidates in preparation for listing for cardiac transplantation. Patient, 64 y. o., with chronic heart failure NYHA III class, had developed a rare complication while performing that procedure – a rupture of segmental branch of pulmonary artery (PA) with pulmonary haemorrhage. The episode of pulmonary bleeding was stopped conservatively without surgical management. There was a pseudoaneurysm formation of segmental branch of PA 2,7 × 2,8 cm with signs of thrombosis. Afterfi ve months the patient underwent heart transplantation without severe complications in perioperative period. No more recurrent episodes of pulmonary haemorrhage were identifi ed.Обязательной процедурой, выполняемой перед включением кандидата на трансплантацию сердца в лист ожидания, является катетеризация правых отделов сердца и легочной артерии (ЛА). У пациентки 64 летс хронической сердечной недостаточностью III функционального класса по NYHA при выполнении данной манипуляции возникло редкое осложнение – разрыв сегментарной ветви ЛА с развитием легочного кровотечения. На фоне комплексной интенсивной терапии эпизод легочного кровотечения был купирован, не потребовав хирургического лечения. Сформировалась ложная аневризма сегментарной ветви ЛА размерами 2,7 × 2,8 см с признаками тромбирования. Спустя пять месяцев пациентка перенесла трансплантацию сердца, периоперационный период протекал без развития тяжелых осложнений, повторных эпизодов легочного кровотечения зафиксировано не было

Будка переводчика-синхрониста: ожидания не всегда оправдываются

This article is aimed at stimulating discussion on pedagogical approaches to interpreter training. It examines certain exceptional situations where professional conference interpreters agree to perform simultaneous interpreting in substandard interpreter booths or other substandard working conditions of varying degrees, e.g. when working at governmental level. Interpretation courses, therefore, should allow time for discussions of such situations and of acceptable scenarios of work in substandard conditions, in order to prepare novice interpreters for real-life interpreting assignments where working conditions are non-negotiable and quitting is not an option to be consideredОбсуждаются исключительные ситуации, когда профессиональные конференц-переводчики соглашаются работать в нестандартных будках или при других ненормальных условиях труда ниже установленного стандарта, что иногда наблюдается, как ни странно, при работе на правительственном уровне. Отмечается, что при обучении конференц- переводчиков необходимо уделять время рассмотрению и обсуждению подобных ситуаций, что позволит полнее подготовить начинающих специалистов к работе в реальных условиях, когда нет возможности обсуждать предложенные условия труда и также нет возможности отказаться от выполнения своих обязанносте

Будка переводчика-синхрониста: ожидания не всегда оправдываются

This article is aimed at stimulating discussion on pedagogical approaches to interpreter training. It examines certain exceptional situations where professional conference interpreters agree to perform simultaneous interpreting in substandard interpreter booths or other substandard working conditions of varying degrees, e.g. when working at governmental level. Interpretation courses, therefore, should allow time for discussions of such situations and of acceptable scenarios of work in substandard conditions, in order to prepare novice interpreters for real-life interpreting assignments where working conditions are non-negotiable and quitting is not an option to be consideredОбсуждаются исключительные ситуации, когда профессиональные конференц-переводчики соглашаются работать в нестандартных будках или при других ненормальных условиях труда ниже установленного стандарта, что иногда наблюдается, как ни странно, при работе на правительственном уровне. Отмечается, что при обучении конференц- переводчиков необходимо уделять время рассмотрению и обсуждению подобных ситуаций, что позволит полнее подготовить начинающих специалистов к работе в реальных условиях, когда нет возможности обсуждать предложенные условия труда и также нет возможности отказаться от выполнения своих обязанносте

Dynamic bending rigidity of a 200-bp DNA in 4 mM ionic strength: a transient polarization grating study.

DNA may exhibit three different kinds of bends: 1) permanent bends; 2) slowly relaxing bends due to fluctuations in a prevailing equilibrium between differently curved secondary conformations; and 3) rapidly relaxing dynamic bends within a single potential-of-mean-force basin. The dynamic bending rigidity (kappa(d)), or equivalently the dynamic persistence length, P(d) = kappa(d)/k(B)T, governs the rapidly relaxing bends, which are responsible for the flexural dynamics of DNA on a short time scale, t < or = 10(-5) s. However, all three kinds of bends contribute to the total equilibrium persistence length, P(tot), according to 1/P(tot) congruent with 1/P(pb) + 1/P(sr) + 1/P(d), where P(pb) is the contribution of the permanent bends and P(sr) is the contribution of the slowly relaxing bends. Both P(d) and P(tot) are determined for the same 200-bp DNA in 4 mM ionic strength by measuring its optical anisotropy, r(t), from 0 to 10 micros. Time-resolved fluorescence polarization anisotropy (FPA) measurements yield r(t) for DNA/ethidium complexes (1 dye/200 bp) from 0 to 120 ns. A new transient polarization grating (TPG) experiment provides r(t) for DNA/methylene blue complexes (1 dye/100 bp) over a much longer time span, from 20 ns to 10 micros. Accurate data in the very tail of the decay enable a model-independent determination of the relaxation time (tau(R)) of the end-over-end tumbling motion, from which P(tot) = 500 A is estimated. The FPA data are used to obtain the best-fit pairs of P(d) and torsion elastic constant (alpha) values that fit those data equally well, and which are used to eliminate alpha as an independent variable. When the relevant theory is fitted to the entire TPG signal (S(t)), the end-over-end rotational diffusion coefficient is fixed at its measured value and alpha is eliminated in favor of P(d). Neither a true minimum in chi-squared nor a satisfactory fit could be obtained for P(d) anywhere in the range 500-5000 A, unless an adjustable amplitude of azimuthal wobble of the methylene blue was admitted. In that case, a well-defined global minimum and a reasonably good fit emerged at P(d) = 2000 A and <deltazeta(2)>(1/2) = 25 degrees. The discrimination against P(d) values <1600 A is very great. By combining the values, P(tot) = 500 A and P(d) = 2000 A with a literature estimate, P(pb) = 1370 A, a value P(sr) = 1300 A is estimated for the contribution of slowly relaxing bends. This value is analyzed in terms of a simple model in which the DNA is divided up into domains containing m bp, each of which experiences an all-or-none equilibrium between a straight and a uniformly curved conformation. With an appropriate estimate of the average bend angle per basepair of the curved conformation, a lower bound estimate, m = 55 bp, is obtained for the domain size of the coherently bent state. Previous measurements suggest that this coherent bend is not directional, or phase-locked, to the azimuthal orientation of the filament

Contemporary treatment of pulmonary arterial hypertension: the North-West Registry data analysis

Aim. Using the prospective Registry data, to assess the effects of conventional and specific therapy on the clinical course and survival of the patients with pulmonary arterial hypertension (PAH). Material and methods. The study included 124 patients (mean age 38,2±13,7 years; 34 men and 78 women): 31 with idiopathic PAH (IPAH), 52 with Eisenmenger syndrome, 17 with inoperable chronic thromboembolic pulmonary hypertension, 9 with PAH and corrected congenital heart disease, 6 with PAH and systemic scleroderma, and 6 with PAH and HIV infection. Results. The cumulative one-year and three-year survival rates were 94% and 75%, respectively. Irrespective of the absence of right heart catheterisation and vasoreactive testing, 42,7% of the patients were treated with calcium antagonists. PAH-specific therapy was administered to 40,3% of the participants (64,5% and 21% of those with IPAH and Eisenmenger syndrome, respectively). PAH-specific therapy was associated with an increase in survival time. Conclusion. In PAH patients, the prognosis is linked to early administration of specific monotherapy and possible combination therapy. Developing a national registry of pulmonary hypertension will facilitate the assessment of the real-world demand for specific therapy and the related costs