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The transfection of SV40- β-gal plasmid was used as the normalized control.
The comparative CT method was used to quantify the expression of SPL, SGPP1 and SGPP2 using GAPDH as the normalized control.
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The relative expression level of each candidate gene was calculated using GAPDH as the internal normalized control with the same calibrator.
The miRNeasy Serum/Plasma Spike-In Control, a Caenorhabditis elegans miR-39 miRNA mimic, was chosen as the normalized internal control.
β-actin was used as a normalized control, using the following primers: 5′-GCA CCC AGC ACA CAA TGA AG-3′ (forward) and 5′-GCA CCC AGC ACA ATG AAG-3′ (reverse).
The endogenous actin gene was used as a normalized control gene.
The housekeeping GAPDH (1 2000, Santa Cruz Biotechnology) was detected as a normalized control.
TaqMan® MicroRNA Assays (Applied Biosystems, CA, USA) was used as the probe for has-miR-195 and has-U6 which act as a normalized control.
USA300 was used as a normalized control to measure sample expression.
Renilla luciferase acted as a reporter gene and firefly luciferase as a normalized control for each individual analysis.
The current amplitude (I) values (current amplitude = integrated current/acquisition time) were then normalized to the corresponding controls obtained from the same patches to yield normalized NPo (control as 1), as the normalized current amplitude was equivalent to the normalized NPo obtained from single-channel analysis when the single-channel conductance remains the same [32].
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