Silencing effects

Nesvijski, E.G., Nogin, S.I. Acoustic Emission Technics for Nondestructive Evaluation of Stress of Concrete and Reinforced Concrete Structures and Materials. Third Conference on Nondestructive Evaluation of Civil Structures and Materials, Boulder, CO, 1996. Nesvijski, E. G. Failure Forecast and the Acoustic Emission Silence Effect in Concrete. ASNT s Spring Conference, Houston, TX, 1997. 

To optimize the amount of siRNA for in vitro transfection, prepare the pep tide/siRNA complexes with various concentrations of siRNA, ranging from 1 pmol to 100 pmol per ml. The optimal amount of siRNA required for efficient gene silencing effects varies between the cell types used. 

Depending on the cell division rate, optimal gene silencing effects may be observed at different time points post transfection. It is suggested to perform western blotting at 48-96 h post transfection to determine the optimal time point. 

Oishi et al. coupled lactosylated PEG to siRNA via an acid labile (S-thiopropio-nate bond, followed by complexation with poly(L-lysine) into polyion complex micelles. The complex was internalized by receptor-mediated endocytosis into hepatoma cells and the gene silencing effect of RNAi was considerably enhanced compared to the free conjugate, demonstrating the effect of pH-sensitive PEGylation [56]. 

This method is suggested as an alternative method for siRNA-lipoplex formation, when the spontaneous formation of lipoplex did not produce an effective gene silencing effect (See Note 13). It is important to note that application of vortex-mixing to form pDNA-lipoplex is not practicable, since it may damage the structure of pDNA. 

Fig. 1. Effect of vortex speed (2,500 rpm for 10 min) on the luciferase gene siiencing efficiency of a siRNA-LipoTrusf lipoplex in HeLa cells. The amount of cationic lipid was 9.6 aM. The siRNA doses correspond to the cationic Npid+/siRNA- charge ratio of 30.45, 15.24,7.62,3.81,1.90 and 0.95, respectively. LipoTrusf is constituted of dioleoylphos-phatidylethanolamine, cholesterol and the cationic lipid 0,0 -ditetradecanoyl-/V-(a-trimethyl ammonioacetyl) diethanolamine chloride (DC-6-14) in the molar ratio of 0.75/0.75/1.00. For this cationic liposome, an expressive gene silencing effect in vitro was obtained at lower siRNA dose with application of a higher vortex-mixing during complex formation...

Du Q, Thonberg H, Wang J, et al. (2005). A systematic analysis of the silencing effects of an active siRNA at all single-nucleotide mismatched target sites. Nucleic Acids Res. 33 1671-1677. 

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