156 research outputs found
Real-time DNA microarray analysis
We present a quantification method for affinity-based
DNA microarrays which is based on the
real-time measurements of hybridization kinetics.
This method, i.e. real-time DNA microarrays,
enhances the detection dynamic range of conventional
systems by being impervious to probe
saturation in the capturing spots, washing
artifacts, microarray spot-to-spot variations, and
other signal amplitude-affecting non-idealities. We
demonstrate in both theory and practice that the
time-constant of target capturing in microarrays,
similar to all affinity-based biosensors, is inversely
proportional to the concentration of the target
analyte, which we subsequently use as the fundamental
parameter to estimate the concentration
of the analytes. Furthermore, to empirically
validate the capabilities of this method in practical
applications, we present a FRET-based assay which
enables the real-time detection in gene expression
DNA microarrays
A BBPāMud2p heterodimer mediates branchpoint recognition and influences splicing substrate abundance in budding yeast
The 3ā² end of mammalian introns is marked by the branchpoint binding protein, SF1, and the U2AF65-U2AF35 heterodimer bound at an adjacent sequence. Baker's yeast has equivalent proteins, branchpoint binding protein (BBP) (SF1) and Mud2p (U2AF65), but lacks an obvious U2AF35 homolog, leaving open the question of whether another protein substitutes during spliceosome assembly. Gel filtration, affinity selection and mass spectrometry were used to show that rather than a U2AF65/U2AF35-like heterodimer, Mud2p forms a complex with BBP without a third (U2AF35-like) factor. Using mutants of MUD2 and BBP, we show that the BBPāMud2p complex bridges partner-specific Prp39p, Mer1p, Clf1p and Smy2p two-hybrid interactions. In addition to inhibiting Mud2p association, the bbpĪ56 mutation impairs splicing, enhances pre-mRNA release from the nucleus, and similar to a mud2::KAN knockout, suppresses a lethal sub2::KAN mutation. Unexpectedly, rather than exacerbating bbpĪ56, the mud2::KAN mutation partially suppresses a pre-mRNA accumulation defect observed with bbpĪ56. We propose that a BBPāMud2p heterodimer binds as a unit to the branchpoint in vivo and serves as a target for the Sub2p-DExD/H-box ATPase and for other splicing factors during spliceosome assembly. In addition, our results suggest the possibility that the Mud2p may enhance the turnover of pre-mRNA with impaired BBP-branchpoint association
Murine and Bovine Ī³Ī“ T Cells Enhance Innate Immunity against Brucella abortus Infections
Ī³Ī“ T cells have been postulated to act as a first line of defense against infectious agents, particularly intracellular pathogens, representing an important link between the innate and adaptive immune responses. Human Ī³Ī“ T cells expand in the blood of brucellosis patients and are active against Brucella in vitro. However, the role of Ī³Ī“ T cells in vivo during experimental brucellosis has not been studied. Here we report TCRĪ“ā/ā mice are more susceptible to B. abortus infection than C57BL/6 mice at one week post-infection as measured by splenic colonization and splenomegaly. An increase in TCRĪ³Ī“ cells was observed in the spleens of B. abortus-infected C57BL/6 mice, which peaked at two weeks post-infection and occurred concomitantly with diminished brucellae. Ī³Ī“ T cells were the major source of IL-17 following infection and also produced IFN-Ī³. Depletion of Ī³Ī“ T cells from C57BL/6, IL-17RĪ±ā/ā, and GMCSFā/ā mice enhanced susceptibility to B. abortus infection although this susceptibility was unaltered in the mutant mice; however, when Ī³Ī“ T cells were depleted from IFN-Ī³ā/ā mice, enhanced susceptibility was observed. Neutralization of Ī³Ī“ T cells in the absence of TNF-Ī± did not further impair immunity. In the absence of TNF-Ī± or Ī³Ī“ T cells, B. abortus-infected mice showed enhanced IFN-Ī³, suggesting that they augmented production to compensate for the loss of Ī³Ī“ T cells and/or TNF-Ī±. While the protective role of Ī³Ī“ T cells was TNF-Ī±-dependent, Ī³Ī“ T cells were not the major source of TNF-Ī± and activation of Ī³Ī“ T cells following B. abortus infection was TNF-Ī±-independent. Additionally, bovine TCRĪ³Ī“ cells were found to respond rapidly to B. abortus infection upon co-culture with autologous macrophages and could impair the intramacrophage replication of B. abortus via IFN-Ī³. Collectively, these results demonstrate Ī³Ī“ T cells are important for early protection to B. abortus infections
Cardiomyocytes Sense Matrix Rigidity through a Combination of Muscle and Non-muscle Myosin Contractions
British Heart Foundatio
Spin Symmetry Requirements in Density Functional Theory: The Proper Way to Predict Magnetic Coupling Constants in Molecules and Solids
Holdup for two-phase flow of gas-non-newtonian liquid mixtures in horizontal and vertical pipes
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