Proposal of a New Hybrid Breeding Method Based on Genotyping, Inter-Pollination, Phenotyping and Paternity Testing of Selected Elite F1 Hybrids.

Testing inbred lines for their combining ability is, due to high numbers of line to line testing needed for determination of hybrid performance, the most limiting factor in the F1 hybrid breeding procedure. We propose a novel method of F1 hybrid breeding that enables evaluation of large number of line to line crosses for their hybrid performance. Inbred lines (preferably doubled haploid - DH) are produced from heterozygous populations, genotyped and maintained. A group of lines is inter-pollinated randomly and their progeny examined. To identify elite F1 hybrids, these individual plants are selected by their superior phenotypic characteristics. Finally using paternity testing only of selected hybrids, the origin of paternal lines is revealed. To predict the number of F1 offspring needed in relation to the number of inbred lines being inter-pollinated, a mathematical formula was developed. For instance, using this formula for the inter-pollination of 60 distinct lines, the probability of obtaining all descendants of paternal-parent lines in a maternal-parent row represented at least once is achieved with 420 F1 plants in a row (p = 0.95). In a practical experiment with white cabbage, DH lines were produced using microspore culture; plants were grown to maturity and genotyped at eight polymorphic SSR loci. Two groups of lines (36 and 33 lines per group) were inter-pollinated by two methods, either using cage pollination with bumblebees or using open pollination in isolated field. A total of 9,858 F1 plants were planted and based on their phenotypic characteristics 213 were selected as elite phenotypes. 99 of them were genetically diverse and 5 of them were selected as super elite. Selected plants were analysed by the same SSR markers and the paternal origin of selected F1 plants was determined. Out of 213 selected elite plants 48 were reciprocals thus exhibiting power of selection based on single plant. We demonstrate that this new approach to hybrid development is efficient in white cabbage and we propose breeders to test it in various vegetable and crop species. Moreover, some other aspects of the proposed technique need to be tested and verified both for practical and economic criteria.

The effect of high frequency electric pulses on muscle contractions and antitumor efficiency in vivo for a potential use in clinical electrochemotherapy.

Muscle contractions present the main source of unpleasant sensations for patients undergoing electrochemotherapy. The contractions are a consequence of high voltage pulse delivery. Relatively low repetition frequency of these pulses (1 Hz) results in separate muscle contractions associated with each single pulse that is delivered. It would be possible to reduce the number of unpleasant sensations by increasing the frequency of electric pulses above the frequency of tetanic contraction, provided that the antitumor efficiency of electrochemotherapy remains the same. These assumptions were investigated in the present paper by measuring the muscle torque at different pulse repetition frequencies and at two different pulse amplitudes in rats and studying the antitumor efficiency of electrochemotherapy at different pulse repetition frequencies on tumors in mice. Measurements of muscle torque confirmed that pulse frequencies above the frequency of tetanic contraction (>100 Hz) reduce the number of individual contractions to a single muscle contraction. Regardless of the pulse amplitude, with increasing pulse frequency muscle torque increases up to the frequency of 100 or 200 Hz and then decreases to a value similar to that after application of a 1 Hz pulse train. Electrochemotherapy in vivo with higher repetition frequencies inhibits tumor growth and is efficient at all pulse frequencies examined (1 Hz-5 kHz). These results suggest that there is a considerable potential for clinical use of high frequency pulses in electrochemotherapy.

Techniques of signal generation required for electropermeabilization. Survey of electropermeabilization devices.

Electropermeabilization is a phenomenon that transiently increases permeability of the cell plasma membrane. In the state of high permeability, the plasma membrane allows ions, small and large molecules to be introduced into the cytoplasm, although the cell plasma membrane represents a considerable barrier for them in its normal state. Besides introduction of various substances to cell cytoplasm, permeabilized cell membrane allows cell fusion or insertion of proteins to the cell membrane. Efficiency of all these applications strongly depends on parameters of electric pulses that are delivered to the treated object using specially developed electrodes and electronic devices--electroporators. In this paper we present and compare most commonly used techniques of signal generation required for electropermeabilization. In addition, we present an overview of commercially available electroporators and electroporation systems that were described in accessible literature.

Three methods for calculation of the hyper-Wiener index of molecular graphs.

The hyper-Wiener index WW of a graph G is defined as WW(G) = (summation operator d (u, v)(2) + summation operator d (u, v))/2, where d (u, v) denotes the distance between the vertices u and v in the graph G and the summations run over all (unordered) pairs of vertices of G. We consider three different methods for calculating the hyper-Wiener index of molecular graphs: the cut method, the method of Hosoya polynomials, and the interpolation method. Along the way we obtain new closed-form expressions for the WW of linear phenylenes, cyclic phenylenes, poly(azulenes), and several families of periodic hexagonal chains. We also verify some previously known (but not mathematically proved) formulas.