Using cannabis DNA as the internal control ensures plant cells have been lysed to release pathogens growing endophytically, to ensure the DNA survived extraction intact, and to ensure there is no contamination in your Assay Master Mix.
Accurate pathogen detection with qPCR requires the use of internal controls to rule out any false negatives. Successfully detecting the internal control gives the lab technician confidence that the test would have detected the target pathogen had it been present in the sample. Without an internal control, the technician wouldn't know whether they missed a well and never added any sample material.
Internal controls also help the lab technician troubleshoot any potential problems that could compromise the qPCR results, such as unsuccessful DNA extraction, PCR inhibition, or cross-contamination.
All internal controls in Medicinal Genomic qPCR assays target a sequence in the cannabis genome. This makes sense because cannabis DNA should be present in any test of cannabis material. Other platforms include an internal control in their master mix, but we believe using a cannabis-based internal control is valuable for 3 reasons.
Cannabis DNA internal controls ensure a clean DNA extraction
Accurately detecting the internal cannabis control indicates the lab technician extracted intact DNA from the sample and inhibitors are not preventing PCR amplification. With Medicinal Genomics kits, you know you've added DNA that is clean and intact when you see Hex signal in all the sample wells. That should give you confidence that if there is no signal from the other channels, that is because there is no target DNA present.
Cannabis DNA internal controls ensure endophytes are properly sampled
Some pathogens live their entire life endophytically. Meaning they gain access to the inside of a plant either through damage to cell walls, or through stomata on leaf surface. If technicians don't lyse open plant cells they might not gain access to the pathogen inside and it can go undetected.
Detecting cannabis DNA in the internal control means the plant cells have been properly lysed, and any endophytes that contain target DNA will also be detected.
Cannabis DNA internal controls ensure no cross-contamination
Each PathoSEEK instrument run includes a positive and negative control for each assay. The positive control well should show signal for the target pathogen, while the negative control well should show no signal at all. This ensures that no cannabis DNA was cross-contaminated when adding samples.
Platforms that include an 'internal control' in the master mix will see signal in every well the master mix is used. that means you will never know if you've cross-contaminated something when adding samples.