14,469 research outputs found
On the Tau-Cycle Condition
AbstractThere is a set of equivalence conditions for the orthonormality of the compactly supported scaling functions. Among them, there is the Cohen's τ-cycle condition. In order to answer the question whether it is enough to check this condition by a finite number of points, we study the τ-cycles in more detail
Cloning and Expression of the Xylanase Gene From Bacillus Coagulans and the M Gene of Newcastle Disease Virus in Lactococcus Lactis
Lactococcus lactis is being developed as a vaccine delivery system as it bears no
threat to animal and human health. It has been used for centuries in the fermentation
of foods and is generally recognised as safe. However, a safety factor pertaining to
the type of selectable marker present on the vector system poses to be of concern.
Chances of the transfer of antibiotic resistance genes, usually employed by vectors as
selectable markers, into the natural environment become a possible risk. Therefore,
there is a need for the development of ideal vectors without such selectable markers
(Dertzbaugh, 1998).
The activity of the xylanase gene of Bacillus coagulans ST -6 can be detected on
Remazol Brilliant Blue-Xylan (RBB-Xylan) as a clear halo zone against a dark blue
background. This characteristic allows xylanase to be used as a selectable
chromogenic marker on any vector system. On the other hand, the matrix or
membrane (M) protein of Newcastle disease virus (NDV) strain AF 2240 can be
useful as an antigen to generate antibody response towards NDV infection in
chickens.Both, the xylanase gene (0.8 kb) of B. coagulans ST-6 and the M gene (1.1 kb) of
NDV strain AF 2240 were cloned in lactococcal expression vector pMG36e and
transformed into Escherichia coli XLI-blue MRF'. The recombinant plasmids,
pMG36e-X and pMG36e-X-M were sub-cloned into L. lactis MG 1363 via
electroporation. The insertion and orientation of the xylanase gene was confmned
using restriction enzyme analysis and PCR amplification. Xylanase activity and
expression on RBB-Xylan agar plates further confmned its presence in the
recombinant plasmid pMG36e-X. In addition, the enzyme activity was also
quantitatively showed using the Somogyi-Nelson assay. The sequence of the M gene
obtained in clone pMG36e-X-M from L. lactis MG 1363 was found to be 99%
homologous to the established sequence (Jemain, 1999). Expression of the fusion M
protein was studied at the transcriptional leve1. RT -PCR was used to detect the
transcription of the gene using RNA of L. lactis MG1363 containing the recombinant
plasmid pMG36e-X-M as template. The size of the RT-PCR product correlated with
the size of the cloned M gene (1.1 kb). In addition, the RT-PCR product was
sequenced to confirm the presence of the M gene.
Based on the results obtained, recombinant plasmids pMG36e-X and pMG36e-X-M
were successfully constructed and introduced into E. coli XLI-blue-MRF' and L.
lactis MG1363. The recombinant DNA pMG36e-X is capable of expressing the
xylanase gene and can be further developed as a chromogenic selection marker for a
new and improved food-grade shuttle vector for E. coli and L. lactis. On the other
hand, expression of the fusion M gene was detected at transcriptional level in
recombinant clone pMG36e-X-M
Tyrosine Phosphorylation linked to Cardiac Arrhythmias via HCN Pacemaker Channel
Cardiac sudden death (CSD) is the leading cause of death in the United States. Although the causes of CSD are not understood completely, cardiac arrhythmias have been indicated as the major contribution to CSD. The main purpose of this thesis is to investigate the mechanisms underlying cardiac arrhythmias, therefore providing scientific basis for clinical applications in the future to lower the rate of CSD.;My thesis contains four chapters. Chapter 1 aims to examine a novel role of tyrosine phosphorylation in the arrhythmogenesis. To achieve this goal, a rat arrhythmia model was established without structural remodeling, and the Src tyrosine kinase and protein tyrosine phosphatase were evaluated in the induction of cardiac arrhythmias by using electrocardiography (ECG) technique (two manuscripts are in preparation).;In chapter 2, we elucidate the mechanism tyrosine phosphorylation used to modulate the cardiac pacemaker channels that directly contribute to the regulation of heart rate and their potential new role in cardiac arrhythmias (J Biol Chem 284: 30433-30440, 2009).;In chapter 3, we explore a novel role of Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels serving as a regulatory protein to regulate the Ca2+ influx via inactivation of L-type calcium channel (LTCC) (Am J Physiol Cell Physiol 298(5): C1029-37, 2010).;In chapter 4, the results that have not been published are summarized. These results represented my efforts to explore previously unrecognized mechanisms that underlie the genesis and development of cardiac arrhythmias that predispose the heart to CSD, claiming more than 300,000 lives each year in the US.;Collectively, our results indicate that tyrosine phosphorylation plays a critical role in the regulation of cardiac rhythm, possibly via modulation the activities of HCN pacemaker channels. Furthermore, we also found that HCN channels can limit the Ca2+ influx via induction of fast inactivation of LTCC. These results not only help us to better understand the mechanisms of cardiac arrhythmias, but can also be applied to the future drug development for the prevention of CSD caused by cardiac arrhythmias
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