In this article we present validation of a real-time RT-PCR method to quantitate mRNA expression levels of atrial natriuretic peptide and c-fos in an in vitro model of cardiac hypertrophy. This method requires minimal sample and no postreaction manipulation. In real-time RT-PCR a dual-labeled fluorescent probe is degraded concomitant with PCR amplification. Input target mRNA levels are correlated with the time (measured in PCR cycles) at which the reporter fluorescent emission increases beyond a threshold level. The use of an oligo(dt) magnetic bead protocol to harvest poly(A) mRNA from cultured cells in 96-well plates minimized DNA contamination. We show that the GAPDH gene chosen for normalization of the RNA load is truly invariant throughout the biological treatments examined. We discuss two methods of calculating fold increase: a standard curve method and the DeltaDelta Ct method. Real-time quantitative RT-PCR was used to determine the time course of c-fos induction and the effect of varying doses of four known hypertrophy agents on atrial naturitic factor messenger RNA expression in cultured cardiac muscle cells. Our results agree with published data obtained from Northern blot analysis.
"The Tth111I polymorphism of the NR3C1 gene was genotyped using the PCR-HRM method with application of LightScanner® 32 System       . Expression of TGFβ1 gene was measured with real-time quantitative PCR (qRT-PCR) method and the 2−ΔΔCT method (ddCT — delta delta cycle threshold values)    . Genotyping and qRT-PCR were performed by two investigators who were unaware of the participants' phenotypes. "
[Show abstract][Hide abstract] ABSTRACT: Personal and environmental factors might have an impact on strategies of coping with stress and temperamental traits according to the Regulative Theory of Temperament in asthmatic patients. They can modify the clinical picture, the course of a disease and effectiveness of treatment. Personal variables are key factors in determining formal characteristic of behavior and effective management method in asthmatic patients.
[Show abstract][Hide abstract] ABSTRACT: Arbuscular mycorrhizal fungi (AMF) are known to improve plants stress tolerance by increasing water absorption and trehalose accumulation. However, the relationship between trehalose and AMF water transport and their role in the mycorrhizal rice water strategy at low temperature has not been established. Different temperature and exogenous trehalose treatment experiments were performed on potted non-mycorrhizal and mycorrhizal rice. The results showed that AMF enhanced rice root water uptake at both normal and low temperatures. At low temperature, the AMF aquaporin (GintAQPF) expression levels were increased and higher rice aquaporin (OsPIPs) expression levels were exhibited in mcyorrhizal rice than non-mycorrhizal rice. The increased trehalose biosynthesis gene transcripts, such as OsTPS1, OsTPS2 and OsTPP1 in mycorrhizal rice roots generated more trehalose than in non-mycorrhizal rice at either normal or low temperature. Application of exogenous trehalose demonstrated that trehalose could regulate AMF and rice water absorption by inducing the expression of GintAQPF and several OsPIPs and create better plant growth conditions. Consequently, we hypothesized that one of the mechanisms by which AMF improve plant resistance to low temperature was AMF-enhanced trehalose accumulation, which could induce AMF and host plants aquaporin expression that then maintain better water relations in mycorrhizal plants at low temperatures.
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