Sentences with phrase «reaction volume»

An average of ∼ 500 ng of total RNA input per reaction was diluted in water to a total volume of 25 µl, and 25 µl of master mix that contained EZ buffer, rTth enzyme, dNTPs, Rox reference dye, MnCl2 and primer / probe was added to obtain a final reaction volume of 50 µl.

For the GAPDH RT - PCR assay, 1.25 µl of TaqMan pre-developed human GAPDH primers (Applied Biosystems) was used in a final reaction volume of 25 µl.

The reaction mix per sample consisted of 6.33 µl H2O, 1 µl Buffer 1, 1 µl Cresol Red 10x, 0.2 µl dNTP mix (10 mM), 0.07 µl polymerase mix, 0.2 µl each forward / reverse primer (25 µM stock) and 1 µl template gDNA for a total reaction volume of 10 µl.

Until now, the application of qPCR in high throughput screening (HTS) has been limited by an often laborious multistep process to generate suitable template, and high reagent cost due to large reaction volumes.

Quantitative PCR was conducted in triplicate in an Applied Biosystems MicroAmp Optical 96 - well Reaction Plate with a 25 µl reaction volume containing 12.5 µl of Thermo Scientific Maxima SYBR Green / ROX qPCR Master Mix, 2 µl of purified DNA and a final primer concentration of 0.15 µM for both forward and reverse primers.

Briefly, 100 ng of RNA was isolated and converted to cDNA in a 50 μl reaction volume using the High - Capacity cDNA Archive Kit (cat # 4322171, Applied Biosystems).

The digestion was performed using BtsI CutSmart (New England BioLabs) with 2.5 µL 10X buffer, 0.40 µL BtsI, 1 µg DNA, and water adjusted accordingly for a total reaction volume of 25 µL.