Effect of DNA Base Modification on Polymerase Chain Reaction Efficiency and Fidelity

Polymerase stop assays, used to quantify DNA damage, assume single lesions are sufficient to block thermostable DNA polymerase progression. To explore this assumption, 90 base oligonucleotides containing normal or modified DNA bases were amplified using real-time PCR. Data implied that the PCR efficiency was influenced to differing degrees depending on which base lesion was present on the input oligonucleotide; specifically, while reactions with templates containing a single 8-oxo-7,8-dihydro-2í-deoxyguanosine (8-oxodG) were not noticeably altered, the presence of a single 8-oxo-7,8-dihydro-2í-deoxyadenosine, an abasic site, or a cis-syn thymidine dimer (TT dimer) dramatically delayed amplification. In addition, the presence of two tandem 8-oxodGs substantially hindered amplification when compared with two 8-oxodGs separated by 13 bases which indicated that the position of lesions also influenced the PCR. To quantify variations in amplification, novel mathematical formulae were developed which report differences in exponential amplification as rates of damage bypass. These treatments assume each template in the PCR is damaged to the same degree. Quantification of damage to cellular DNA, which is a mixture of damaged and undamaged template, required further refinement of real-time PCR mathematics; differences in amplification were defined in terms of damage probability (lesion frequency) rather than lesion bypass rate. The validity of these formulae was determined using DNA samples quantified previously using current polymerase stop methods. In addition to impacting reaction efficiency, DNA base modifications decreased reaction fidelity. In reactions with templates containing 8-oxodGs, both the normal Watson/Crick association with dCMP as well as the incorporation of dAMP occurred at the lesion site. Despite similar structural characteristics, the existence of 8-oxodA resulted in a pronounced n-1 deletion in addition to the normal association with dTMP. Sequence data from abasic and TT dimer modifications were inconclusive but suggested the presence of multiple nucleotide incorporation events opposite the modifications. The present work enabled the adaptation of real-time PCR for DNA damage quantification, identified DNA base lesions as potential PCR mutagens, and provides the basis for further refinement of polymerase stop assays as research and clinical tools to monitor DNA damage and repair

Parallelization of sequential programs: distribution of arrays among processors and structurization of communications

Data distribution functions are introduced. They are matced with scheduling functions. The processors and iterations are determined that use an array element at its fixed position in a statement. This makes it possible to obtain the initial data distribution and also information on the data volume for every processor and on the structure of required communication

Explicit expressions for meromorphic solution of autonomous nonlinear ordinary differential equations

Meromorphic solutions of autonomous nonlinear ordinary differential equations are studied. An algorithm for constructing meromorphic solutions in explicit form is presented. General expressions for meromorphic solutions (including rational, periodic, elliptic) are found for a wide class of autonomous nonlinear ordinary differential equations

Driven electronic bridge processes via defect states in 229^{229}Th-doped crystals

The electronic defect states resulting from doping $^{229}$Th in CaF$_2$ offer a unique opportunity to excite the nuclear isomeric state $^{229m}$Th at approximately 8 eV via electronic bridge mechanisms. We consider bridge schemes involving stimulated emission and absorption using an optical laser. The role of different multipole contributions, both for the emitted or absorbed photon and nuclear transition, to the total bridge rates are investigated theoretically. We show that the electric dipole component is dominant for the electronic bridge photon. In contradistinction, the electric quadrupole channel of the $^{229}$Th isomeric transition plays the dominant role for the bridge processes presented. The driven bridge rates are discussed in the context of background signals in the crystal environment and of implementation methods. We show that inverse electronic bridge processes quenching the isomeric state population can improve the performance of a solid-state nuclear clock based on $^{229m}$Th

Enantioselective recognition of amino acids by enantiomerically pure calix[4]arene carboxylic acid or their diastereomerically pure N-(1-phenyl)ethyl amides

The interaction of inherently chiral calix[4]arene carboxilic acids and their amides with amino acids in the organic phase has been studied using electron spectroscopy. It was found that the chiral calix[4]arenes are able of enantioselective recognition of L- and D-forms of amino acids. Stability constants of the calixarene - amino acid supramolecular complexes were determined and mechanism of the host-guest interaction was examined by molecular modeling method. © ISUCT Publishing

NMR spectroscopic detection of chirality and enantiopurity in referenced systems without formation of diastereomers

Enantiomeric excess of chiral compounds is a key parameter that determines their activity or therapeutic action. The current paradigm for rapid measurement of enantiomeric excess using NMR is based on the formation of diastereomeric complexes between the chiral analyte and a chiral resolving agent, leading to (at least) two species with no symmetry relationship. Here we report an effective method of enantiomeric excess determination using a symmetrical achiral molecule as the resolving agent, which is based on the complexation with analyte (in the fast exchange regime) without the formation of diastereomers. The use of N,N′-disubstituted oxoporphyrinogen as the resolving agent makes this novel method extremely versatile, and appropriate for various chiral analytes including carboxylic acids, esters, alcohols and protected amino acids using the same achiral molecule. The model of sensing mechanism exhibits a fundamental linear response between enantiomeric excess and the observed magnitude of induced chemical shift non-equivalence in the 1H NMR spectra

High-speed fixed-target serial virus crystallography

We report a method for serial X-ray crystallography at X-ray free-electron lasers (XFELs), which allows for full use of the current 120-Hz repetition rate of the Linear Coherent Light Source (LCLS). Using a micropatterned silicon chip in combination with the high-speed Roadrunner goniometer for sample delivery, we were able to determine the crystal structures of the picornavirus bovine enterovirus 2 (BEV2) and the cytoplasmic polyhedrosis virus type 18 polyhedrin, with total data collection times of less than 14 and 10 min, respectively. Our method requires only micrograms of sample and should therefore broaden the applicability of serial femtosecond crystallography to challenging projects for which only limited sample amounts are available. By synchronizing the sample exchange to the XFEL repetition rate, our method allows for most efficient use of the limited beam time available at XFELs and should enable a substantial increase in sample throughput at these facilities

Investigation into the annotation of protocol sequencing steps in the sequence read archive

BACKGROUND: The workflow for the production of high-throughput sequencing data from nucleic acid samples is complex. There are a series of protocol steps to be followed in the preparation of samples for next-generation sequencing. The quantification of bias in a number of protocol steps, namely DNA fractionation, blunting, phosphorylation, adapter ligation and library enrichment, remains to be determined. RESULTS: We examined the experimental metadata of the public repository Sequence Read Archive (SRA) in order to ascertain the level of annotation of important sequencing steps in submissions to the database. Using SQL relational database queries (using the SRAdb SQLite database generated by the Bioconductor consortium) to search for keywords commonly occurring in key preparatory protocol steps partitioned over studies, we found that 7.10%, 5.84% and 7.57% of all records (fragmentation, ligation and enrichment, respectively), had at least one keyword corresponding to one of the three protocol steps. Only 4.06% of all records, partitioned over studies, had keywords for all three steps in the protocol (5.58% of all SRA records). CONCLUSIONS: The current level of annotation in the SRA inhibits systematic studies of bias due to these protocol steps. Downstream from this, meta-analyses and comparative studies based on these data will have a source of bias that cannot be quantified at present

ОСНОВНЫЕ ИТОГИ РОССИЙСКО-БЕЛОРУССКОГО СОТРУДНИЧЕСТВА ПО СЕЛЕКЦИИ ОВОЩНЫХ БОБОВЫХ И ЛУКОВЫХ КУЛЬТУР

The Russian-Belarusian cooperation in breeding of legumes and onion crops has resulted in development of new cultivars of pea (Samorodok), bean (Phaeton and Mignon), onion (Palesskaya znahodka, Patrida and Vermeles), winter garlic (Dubkovsky Asilak), which were included in 2014 in the State Register of the Republic of Belarus.Результатом Российско-Белорусского сотрудничества по селекции овощных бобовых и луковых культур стало создание семи совместных урожайных сортов: гороха овощного – Самородок, фасоли – Фаэтон и Мигнон; лука репчатого – Палесская знаходка, Патрыда и Вермелес; чеснока озимого – Дубковский Асилак, внесенных в 2014 году в Государственный реестр Республики Беларусь