Enhancing quality assurance of medical marijuana using DNA fingerprinting

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Marijuana (Cannabis sativa L.) is increasingly being recognized for its properties in treating a number of human diseases, such as multiple sclerosis, chronic pain, seizures and PTSD. Marijuana plants may be initiated from seeds but more commonly are propagated from vegetative cuttings originating from stock plants (mother plants). Seeds are used when a new genetic line (strain) is being considered, but is subject to variation in phenotypic characteristics in the offspring due to segregation. It is not uncommon for a potential mix-up of genetic lines to occur during commercial production, or for seeds or vegetative propagation to give rise to unusual phenotypes. This creates a problem for quality assurance as well as for monitoring a particular line for genetic stability over time. The availability of a DNA fingerprinting method that can be used to distinguish among selected lines, as well as to study the extent of genetic variation among a wide range of lines and seed sources, would be invaluable as a rapid screen for quality assurance and genetic fidelity. In the preceding Engage grant, a set of 7 ISSR primers was identified that produced distinct banding patterns when used with a broad range of DNA sources of Cannabis. This primer set showed that a significant level of genetic variation exists amongst 40 strains of medical marijuana, something not previously reported. An additional set of primers (SFU1, SFU2) was developed that could distinguish male from female plants of marijuana. The objectives of this research are to: 1) Determine the variables affecting the utility of a plant sex testing kit based on primers SFU1-SFU2 developed in the previous work; 2) Continue to evaluate the set of 7 ISSR primers for suitability to develop a DNA fingerprinting test kit that could be utilized commercially to identify select marijuana strains; 3) Assess the conditions that can influence the variability of results in objectives (1) and (2) above; 4) Conduct DNA sequencing to verify the relationships among the different marijuana strains, as well as between female and male plants, and compare the results to the Cannabis genome sequence available on-line and Genbank sequences.