Thirty primers for the selected gene candidates and for the internal and negative controls were added in triplicate to 96-well plates, and the samples were measured using one plate per sample.
In addition, PCR products of chickpea Actin cDNA [GenBank: AJ012685] and Neomycin phophotransferase (NPTII) gene from the vector pCAMBIA 1305.1 [GenBank: AF354045] were spotted as internal and negative controls respectively to normalize the signals of two replicate blots corresponding to stressed/unstressed chickpea cultivars and to subtract the background intensity respectively.
For each gene, three repeated experiments, including internal controls and negative controls (reaction samples without cDNA templates), were conducted.
It provides explicit comparisons with internal positive and negative controls, thus reducing the impact of systematic or experimental errors.
Furthermore, we also used islet of Langerhans and nontumor ducts present in whole tissue sections as internal positive and negative controls, respectively, to assess staining quality.
Internal positive and negative controls, including normal squamous epithelium of the esophagus from non-cancer patients were utilized as available to further support the staining patterns.
For quality control, two control strains of Escherichia coli (ATCC 25922 and 35218) were tested as well as the internal positive and negative controls included in each microdilution plate.
We will monitor quality control of laboratory tests by ensuring internal positive and negative controls are within the parameters of the test kit for each assay and by evaluating for trends over time.
A negative internal control signal was accepted in case of a positive target sequence with correct positive and negative control signals.
This is a useful model as the fluorescence of the EGFR-GFP served as a convenient measure of EGFR expression and the partial transfection created internal EGFR positive and negative controls.
