Biotechnology Tools to Grow Cannabis

Table of Contents

Cannabis is a multipurpose crop, in which the scientific community is very much interested to harness its power and values for economical uses.

Overview

Cannabis is a multipurpose crop, in which the scientific community is very much interested to harness its power and values for economical uses.

Since ancient times, cannabis has been known for its medicinal and textile uses. The plant is a rich source of fibers, oils, secondary molecules, and other phytochemicals. Additionally, it has some wonderful agricultural features including good resistance to drought and pests, lower water requirements than other plants, such as cotton, and a well-developed root system to prevent soil erosion.

This great versatility of the crop has made it popular among consumers, culturists, and researchers. After its legalization in many American states and some other countries, it has opened the door to explore and dig deep into the possibilities of the uses of plants for medicinal or industrial uses.

In this article, we will review what are the current propagation techniques available for producing cannabis at a commercial scale, and then we’ll learn in brief about the most advanced biotechnology tools developed to overcome the limitation of currently available techniques.

Techniques for Cannabis Propagation

Traditionally, cannabis is grown using seeds or by vegetative propagation, using stem cutting. However, these methods have several limitations including:

• Genetic instability
• Requirement of a longer period for plant maturity
• Less reproducibility
• Prone to diseases and infections

The challenge is resolved by using the tissue culture technique, which is an advanced method to grow plants. In this technique, plants are grown in labs under completely artificial conditions using only a few tissues of plants that are called explants. The advantages offered by these techniques over traditional approaches include:

• High multiplication rate
• Production of disease-free plants
• Less/no variation
• Selective cloning

The most popular and well-studied tissue culture techniques are cell suspension culture, thin cell layer culture, and hairy root cultures.

The success of these techniques depends on several factors including the composition of media, age, type, and size of explants used for the propagation, health of the mother plant, carbohydrate sources, and culture conditions.

The tissue culture technique is especially effective when you are targeting to increase the production of cannabidiols, achieve plants with low or no THC (psychedelic drug, which is if present more than 0.3% in plants makes it illegal for its use in developing any products), achieve disease-free plants, and produce bioengineered biomolecules.

Genetic Engineering of Cannabis Plants

In recent years genetic tools have been proven to be an effective technique in improving crop varieties. They are efficient for improving crop genetics as well. And, combining these tools with tissue culture techniques is another effective approach that’s in use to improve the characteristics of plants and their productivity.

Genetic transformation techniques all introduce certain specific genes with important traits such as herbicide resistance, insect resistance, and pro-vitamin A production.

Given below are some efforts made in developing biotechnology tools for cannabis propagation:

• To study the function of genes in cannabis plants, techniques like targeting-induced local lesions in genomes (TILLING) and virus-induced gene silencing (VIGS) approaches have been used.
• Some applicable techniques to cannabis are Agrobacterium- and Biolistic-mediated gene transformation systems, de novo meristem induction, and virus-assisted gene editing. Among all these techniques biolistic-mediated gene transformation has not been tested on cannabis and the agrobacterium-mediated gene transfer is directly dependent on tissue culture.

Agrobacterium-mediated gene transfer

Agrobacterium tumefaciens is a soil-plant pathogen, which causes crown gall diseases by inserting its pathogenic segment of DNA into plants. Scientists utilize the mechanism of this bacteria to transfer some essential traits into plants. This is done by placing the pathogenic part of T-DNA with the desired genes.

This technique has great attention and interest from scientists for gene transformation of cannabis due to the susceptibility of cannabis genes towards Agrobacterium. However, it also has certain limitations, which includes:

• low efficiency of gene transformation
• low rates of regeneration
• chimeric regeneration including both non-transgenic and transgenic cells and tissues
• inactivation of the transgene

Figure: An illustration of Agrobacterium-mediated Cannabis transformation

CRISPR-Cas-Mediated genome editing

CRISPR-Cas is a tool discovered in bacterial species, where it functions to protect the host organism from pathogenic nucleic acids by specifically destroying them. This genome editing tool has exceptionally improved the plant biotechnology area. It provides a sustainable approach to improving cannabis varieties with desired traits. Normally, the success of the CRISPR-Cas system in developing transgenic crops depends on the selection of the best guide sequence. However, this is still an ongoing research area.

The challenge in designing the candidate gRNA is the non-availability and accessibility of a high-quality reference cannabis genome. Though the cannabis genome has been assembled de novo over a decade ago, however, there are 12 different genomes reported with significant differences in reported genome size, chromosome order, and gene annotations among cannabis genome assemblies, which makes it difficult to design a right gRNA.

Therefore, even though the use of CRISPR-Cas technology in developing transgenic plants is potentially effective, it has its own challenges that hinder its direct application in cannabis propagation.

Figure: An illustration of Cannabis genome engineering techniques.

Choose Plant Cell Technology to Enhance Your Cannabis Tissue Culture Experience

To help the cannabis culturists with their tissue culture processes and help them to pass through the challenging phases, PCT is providing world-class consulting services.

The consulting services are available in two forms: one-on-one phone calls and on-site visits. So, if you need an instant solution for your specific challenges you can have a one-on-one call with our scientists and get your answers instantly.

However, if you are someone building a cannabis tissue culture lab or are already an established lab that is looking to train its staff or wants solutions for pathogen eradication in its commercial-scale plants, you can choose an on-site visit consulting service. Our team will visit your lab and will guide you through the whole process.

The services included in the on-site visits are:

• Blueprint, Budget, & Equipment: build and operate a successful tissue culture lab.Media Preparation: 2 sets of proprietary media preparation SOPs (4 in vitro shoot multiplication protocols and 5 in vitro rooting protocols along with coaching on how to conduct factorial trials).
• Micropropagation: how to select, surface sterilize, and induce nodes into media for removal of surface pathogens and in vitro cloning applications.
• Meristem Dissection: how to dissect apical meristem to remove viruses including HpLvd, Cannabis Cryptic Virus, Lettuce Chlorosis Virus, and more.
• Synthetic Seed and Cryopreservation: Long term genetic storage solutions.
• Pathogen Remediation by Media Amendments: how to remove viruses, systemic fungi, bacterial infections, and endophytes.
• Gender and Pathogen Screening by PCR: Eliminate male plants, identify cannabinoid ratios, and protect your mothers from pathogens and pests

Reference

1. Hesami, M., Baiton, A., Alizadeh, M., Pepe, M., Torkamaneh, D., & Jones, A. (2021). Advances and Perspectives in Tissue Culture and Genetic Engineering of Cannabis. International journal of molecular sciences, 22(11), 5671. https://doi.org/10.3390/ijms22115671.
2. Hau-Hsuan Hwang, Manda Yu, and Erh-Min Lai "Agrobacterium-mediated Plant Transformation: Biology and Applications," The Arabidopsis Book 2017(15), (20 October 2017). https://doi.org/10.1199/tab.0186