Simplified 5-minute Leaf-Punch for Accessible Rapid Results of HLVd in Cannabis Plants

By Maria Mcintyre at BioMerieux

INTRODUCTION

The quickly expanding recreational Cannabis industry in the United States and beyond presents unique challenges for production quality from both public health and agricultural standpoints. Accessible rapid diagnostic assays beyond detection of human pathogens to ensure efficient operations are becoming more critical in this growing market. To support these needs here we introduce a novel, streamlined leaf-punch sample preparation methodology for two new GENE-UP assay systems – GENE-UP Gender ID to quickly discriminate male and female plants, and GENE-UP HLVd for the rapid detection of Hop Latent Viroid plant infection.

METHODS

To examine performance of the GENE-UP streamlined cannabis leaf sample preparation method, seven leaf samples were sourced for sex determination based on phenotypic (visual) expression of gender and 15 leaf samples were also independently sourced from both suspected HLVd infected and healthy plants  (based on observed typical/atypical plant growth). Each sample was subsequently punched with a 1.2 mm leaf punch (Uni-Core, 1.2 mm punch), and leaf samples were directly introduced into GU Lysis tubes for DNA extraction and PCR conditioning. After a five minute vortex at 2200 RPM, 5.0 uL of each lysate was transferred to a thawed GENE-UP PRO Gender ID or HLVd assay for analysis. Samples were amplified under each assay’s requisite PCR program, and analyzed on the GENE-UP thermal cycler software.

RESULTS

Each sample tested for the GENDER ID and HLVd matched expected results  (observed female or male phenotype and suspected HLVd infection), with an assay turnaround time of less than three hours. Each internal assay control, monitoring for DNA extraction efficiency, polymerase and reverse transcriptase function, demonstrated successful assay performance in each sample. The data supports the robust performance of this streamlined, five minute mechanical lysis sample preparation procedure for the rapid analysis of sex determination and HLVd infection in cannabis plants.