Latex bead immobilisation in PDMS matrix for the detection of p53 gene point mutation and anti-HIV-1 capsid protein antibodies

Two diagnostic chemiluminescent biochips were developed for either the detection of p53 gene point mutation or the serological detection of anti-HIV-1 p24 capsid protein. Both biochips were composed of 24 microarrays of latex beads spots (4×4) (150 m in diameter, 800 m spacing) entrapped in a poly(dimethylsiloxane) elastomer (PDMS). The latex beads, bearing oligonucleotide sequences or capsid protein, were spotted with a conventional piezoelectric spotter and subsequently transferred at the PDMS interface. The electron microscopy observation of the biochips showed how homogeneous and well distributed the spots could be. Point mutation detection on the codon 273 of the p53 gene was performed on the basis of the melting temperature difference between the perfect match sequence and the one base pair mismatch sequence. The hybridisation of a 20-mer oligonucleotide form the codon 273 including a one base pair mutation in its sequence on a biochip arrayed with non-muted and the muted complementary sequences, enabled a clear discrimination at 56°C between muted and wild sequences. Moreover, the quantitative measurement of the amount of muted sequence in a sample was possible in the range 0.4–4 pmol. Serological measurement of anti-HIV-1 p24 capsid protein on the biochip, prepared with 1-m-diameter latex beads, enabled the detection of monoclonal antibodies in the range 1.55–775 ng mL^–1. Such a range could be lowered to 0.775 ng mL^–1 when using 50-nm-diameter beads, which generated a higher specific surface. The validation of the biochip for the detection of anti-HIV-1 capsid protein antibodies was performed in human sera from seropositive and seronegative patients. The positivity of the sera was easily discriminated at serum dilutions below 1:1,000.     

Identités Multilinéaires de Degré 4 pour les Algèbres de Bernstein et de Mutation Non Commutatives

We give the identities generating the spaces of degree 4 multilinear identities satisfied by the non commutative 0th-order Bernstein algebras and the non commutative mutation algebras. For it, we use a method reducing the search for multilinear identities to the resolution of linear systems.     

Nonlinear Phase Shift by Parametric Amplification in Leaky Waveguides

In this article, the cascaded second-order nonlinear phase shifts induced in the signal wave are studied using the three-wave parametric interaction in a four-layered QPM leaky waveguide configuration in Z-cut LiNbO 3 . In this interaction, the pump-andsignal waves are guided modes and the idler is radiated into the substrate. We have analyzed the nonlinear phase shifts induced in the signal wave due to cascaded second-order effects, as a function of different waveguide parameters. These studies should find application in realizing practical all-optical signal processing devices.     

Deamination of the Radical Cation of the Base Moiety of 2′‐Deoxycytidine: A Theoretical Study

Five pathways leading to the deamination of cytosine (to uracil) after formation of its deprotonated radical cation are investigated in the gas phase, at the UB3LYP/6‐311G(d,p) level of theory, and in bulk aqueous solvent. The most favorable pathway involves hydrogen‐atom transfer from a water molecule to the N3 nitrogen of the deprotonated radical cation, followed by addition of the resulting hydroxyl radical to the C4 carbon of the cytosine derivative. Following protonation of the amino group (N4), the C4? N4 bond is broken with elimination of the NH₃^?+ radical and formation of a protonated uracil. The rate‐determining step of this mechanism is hydrogen‐atom transfer from a water molecule to the cytosine derivative. The associated free energy barrier is 70.2 kJ mol^?1.     

A MELAS syndrome family harboring two mutations in mitochondrial genome

Mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) syndrome is a genetically heterogeneous mitochondrial disorder with variable clinical symptoms. Here, from the sequencing of the entire mitochondrial genome, we report a Korean MELAS family harboring two homoplasmic missense mutations, which were reported 9957T>C (Phe251Leu) transition mutation in the cytochrome c oxidase subunit 3 (COX3) gene and a novel 13849A>C (Asn505His) transversion mutation in the NADH dehydrogenase subunit 5 (ND5) gene. Neither of these mutations was found in 205 normal controls. Both mutations were identified from the proband and his mother, but not his father. The patients showed cataract symptom in addition to MELAS phenotype. We believe that the 9957T>C mutation is pathogenic, however, the 13849A>C mutation is of unclear significance. It is likely that the 13849A>C mutation might function as the secondary mutation which increase the expressivity of overlapping phenotypes of MELAS and cataract. This study also demonstrates the importance of full sequencing of mtDNA for the molecular genetic understanding of mitochondrial disorders.     

Quivers with potentials and their representations I: Mutations

We study quivers with relations given by noncommutative analogs of Jacobian ideals in the complete path algebra. This framework allows us to give a representation-theoretic interpretation of quiver mutations at arbitrary vertices. This gives a far-reaching generalization of Bernstein–Gelfand–Ponomarev reflection functors. The motivations for this work come from several sources: superpotentials in physics, Calabi–Yau algebras, cluster algebras.      

Role of the Residue Q1919 in Increasing Kinase Activity of G2019S LRRK2 Kinase: A Computational Study

Mutations in multi-domain leucine-rich repeat kinase 2 (LRRK2) have been an interest to researchers as these mutations are associated with Parkinson's disease. G2019S mutation in LRRK2 kinase domain leads to the formation of additional hydrogen bonds by S2019 which results in stabilization of the active state of the kinase, thereby increasing kinase activity. Two additional hydrogen bonds of S2019 are reported separately. Here, a mechanistic picture of the formation of additional hydrogen bonds of S2019 with Q1919 (also with E1920) is presented using ‘active’ Roco4 kinase as a homology model and its relationship with the stabilization of the ‘active’ G2019S LRRK2 kinase. A conformational flipping of residue Q1919 was found which helped to form stable hydrogen bond with S2019 and made ‘active’ state more stable in G2019S LRRK2. Two different states were found within the ‘active’ kinase with respect to the conformational change (flipping) in Q1919. Two doubly-mutated systems, G2019S/Q1919A and G2019S/E1920 K, were studied separately to check the effect of Q1919 and E1920. For both cases, the stable S2 state was not formed, leading to a decrease in kinase activity. These results indicate that both the additional hydrogen bonds of S2019 (with Q1919 and E1920) are necessary to stabilize the active G2019S LRRK2.     

A Trait‐based framework for mutation bias as a driver of long‐term evolutionary trends

Previous work has shown that mutation bias can direct evolutionary trends in genotypic space under strong selection and rare mutation. We present an extension of this work to general traits of the organism. We do this by allowing many different genotypes, with different fitnesses, to have the same trait value. This approach makes novel predictions and shows that the outcome of evolution for a trait is influenced by mutation bias as well as the fitness distribution of the genotypes that have the same trait value. This distribution can alter evolution in interesting ways, depending on the likelihood of generating high fitness mutants. We also show that mutation bias can direct evolution when many mutants are present at any one time. We demonstrate that mutation bias can drive long‐term evolutionary trends when the environment is constantly changing. Under biologically realistic conditions, we show that mutation bias can counter strong gradients of environmental selection over time. We conclude that evolutionary trends can be quite independent of the environment, even when they depress population fitness. Finally, we show that entropy can be a powerful source of mutation bias and can drive evolutionary trends. © 2015 Wiley Periodicals, Inc. Complexity 21: 331–345, 2016     

A study of comparability in amplified fragment length polymorphism profiling using a simple model system

A simple amplified fragment length polymorphism (AFLP) model, using the bacteriophage lambda genome, was developed to test the reproducibility of this technique in an international comparative study. Using either non-selective or selective primers, nine fragments or subsets of two or three fragments, respectively, were predicted using in silico software. Under optimized conditions, all predicted fragments were experimentally generated. The reproducibility of the AFLP model was tested by submitting both "unknown" DNA template that had been restricted and ligated with AFLP linkers (R/L mixture) and corresponding primer pairs to nine laboratories participating in the study. Participants completed the final PCR step and then used either slab gel electrophoresis or CE to detect the AFLP fragments. The predicted fragments were identified by the majority of participants with size estimates consistently up to 3 base pair (bp) larger for slab gel electrophoresis than for CE. Shadow fragments, 3 bp larger than the predicted fragments, were often observed by study participants and organizers. The nine AFLP fragments exhibited relative intensities ranging from less than 3% to 22% and, apart from the two weakest fragments, with a % CV of 16 to 25. Fragments containing the highest guanine-cytosine (GC) content of 50-56% showed the greatest stability in the AFLP profiles.     

A novel one cycle allele specific primer extension--molecular beacon displacement method for DNA point mutation detection with improved specificity

We report here a new method for the real-time detection of DNA point mutations with molecular beacon as the fluorescence tracer and 3′ (exo-) Bst DNA polymerase large fragment as the polymerase. The method is based on the mechanism of allele specific primer extension-strand displacement (ASPE-SD). To improve the specificity of the method only one cycle of the allele specific polymerase chain reaction (PCR) was used that could largely eliminate the non-specific reactions between the primers and template of the “wrong” genotype. At first, the primer and molecular beacon both hybridize to the DNA template, and the molecular beacon emits intensive fluorescence. The role of 3′ exonuclease excision of Bst DNA polymerase large fragment is utilized for primer extension. When 3′-termini matches its corresponding template, the primer would efficiently extend and replace the molecular beacon that would simultaneously return to its closed form leading to the quenching of the fluorescence. However, when 3′-termini of the primer mismatches its corresponding template primer extension and molecular beacon displacement would not happen and fluorescence of the hybridized molecular beacon holds the line without fluorescence quenching. This approach was fully demonstrated in synthetic template systems and applied to detect point mutation at codon 259, a possible point mutation site in exon 7 of p53 gene, obtained from human genomic DNA samples with unambiguous differentiation power.