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Plant Breeding and Marker-Assisted Selection: Spotlight on Hemp Cannabis sativa

Read our blog covering recent release of PACE SNP markers in Cannabis sativa for Autoflowering, cannabinoid chemotype and sex determination.

SNP Genotyping with Allele-Specific PCR and PACE Genotyping

SNP markers and SNP genotyping are now widely used in breed improvement programs due to their high-throughput capability, accuracy, and efficiency in marker-assisted selection; PACE technology is ideal for developing, validating and running these breeder-friendly SNP and Indel markers. For further insights into 3CR Bio and Plant Breeding, click here.

Understanding Cannabis: Unraveling its Potential

Cannabis, a versatile crop with multifaceted utility, has historically served myriad purposes, from providing valuable fibers for ropes and nets to yielding protein- and oil-rich seeds. Furthermore, its applications in traditional medicine, dating back to approximately 8000 BCE, and its psychoactive properties underscore its significance in various cultural contexts.

However, despite its versatility, Cannabis is often legally categorized based on the concentration of a single cannabinoid—Δ9-tetrahydrocannabinol (THC). In regions like Canada, the USA, and Europe, Cannabis plants are classified as either hemp-type (with less than 0.3% total THC) or drug-type (with greater than 0.3% total THC). This regulatory framework underscores the need for precise understanding and control of Cannabis genetics to meet legal standards and consumer demands.

Case Study: Understanding the Genetics of Important Traits in Hemp to Help Breed Improved Cultivars

PACE genotyping has already proven itself as an invaluable tool for Cannabis breeders and researchers. Hemp (Cannabis sativa) is an extraordinarily versatile crop, with applications ranging from medicinal compounds to seed oil and fiber products. 3CR Bio spoke with Dr. Jacob Toth, a Plant Breeding and Genetics Post-Doc at Cornell University’s School of Integrated Plant Science, who is part of Professor Larry Smart’s research group. This conversation is featured in our case study  ‘Understanding the genetics of important traits in hemp to help breed improved cultivars.’ We delved into their research and explored how they incorporate PACE® into their workflow.

Cannabis plant breeding
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Unlocking Potential: Open Access Genetic Markers for Sex and Cannabinoid Chemotype in Cannabis sativa

In the realm of hemp breeding and cultivation, the quest for efficient selection and breeding tools has reached a milestone with the development and validation of genetic markers for sex and cannabinoid chemotype. These markers, crafted through high-throughput PACE Genotyping Assays (Allele-specific PCR), signify a pivotal advancement, empowering breeders with precision and efficiency in selecting desired traits. Click here to read the full research paper.

Crucial Traits: Sex and Cannabinoid Chemotype

Sex determination and cannabinoid chemotype stand as pivotal qualitative traits for hemp producers and breeders. The legal landscape demands that hemp cultivars produce less than 0.3% THC by dry weight, underscoring the necessity for understanding genetic predisposition towards THC production. Yet, phenotyping these traits in young plants poses formidable challenges. Until flowering commences, distinguishing between male and female plants remains elusive, compounded by the scant cannabinoid production in immature plants.

Validation and Reliability

The validation of the sex assay across a diverse germplasm spectrum heralds a breakthrough, effectively discerning male plants from their female and monoecious counterparts. Leveraging publicly available sequence data, the development of reliable and high-throughput PACE Genotyping Assays as SNP markers has revolutionized the prediction of sex and cannabinoid chemotype phenotypes in Cannabis sativa. Notably, the transparency of the assay designs, freely accessible for adoption, epitomizes collaboration and progress within the scientific community.

PACE Genotyping: A Game-Changing Marker System

PACE genotyping, with its high-throughput fluorescence-based marker system, emerges as a beacon of innovation, enabling the interrogation of SNPs, indels, and other polymorphic DNA features. Its superiority over gel-based systems is evident, boasting a staggering 45-fold increase in speed. Moreover, PACE’s cost-effectiveness and retention of simple codominance set it apart from other marker systems. Requiring only a qPCR instrument or thermal cycler and a fluorescent plate reader for scoring, the ease and rapidity of PACE genotyping renders this technology tailor-made for breeding endeavours and advanced production systems.

The development and validation of genetic markers for sex and cannabinoid chemotype in Cannabis sativa, propelled by PACE Genotyping Assays and reagents as SNP markers, signify a transformative leap forward in hemp breeding and cultivation. These markers serve as catalysts for innovation, empowering breeders with efficient tools to navigate the intricate landscape of trait selection and cultivation. As we harness the power of genetics to propel hemp cultivation into the future, these markers stand as beacons of progress, guiding the development of improved cultivars tailored to diverse applications and industry needs

Reference: Toth, Jacob A., et al. “Development and validation of genetic markers for sex and cannabinoid chemotype in Cannabis sativa L.” Gcb Bioenergy 12.3 (2020): 213-222.

Unraveling the Potential of Autoflowering Cannabis: A Journey of Innovation

Autoflowering cannabis, once a niche trait, has now become indispensable to breeders and growers alike. This revolutionary feature, driven by genetic innovation, offers a multitude of advantages, propelling the cannabis industry forward and meeting the dynamic demands of consumers worldwide.

The Significance of Autoflowering Traits

Shortened Growth Cycle: Unlike traditional cannabis strains dependent on light cycles, autoflowering varieties bloom based on age, enabling quicker harvests. This attribute proves invaluable, particularly for outdoor cultivators facing limited growing seasons or inadequate sunlight.

Flexibility: Autoflowering plants thrive in diverse environments, including indoor setups where light management can be challenging. This adaptability opens doors for growers to experiment with cultivation techniques and optimize their yields.

Continuous Harvests: By flowering independently of light cycles, autoflowering plants offer growers the flexibility to stagger their planting, ensuring a steady stream of harvests throughout the year. This consistent supply is essential for meeting market demands and maximizing productivity.

Compact Size: Characterized by their smaller stature, autoflowering strains are ideal for discreet or confined growing spaces. This compactness not only enhances cultivation efficiency but also facilitates stealthy operations where privacy is paramount.

Genetic Innovation: Advancements in autoflowering genetics pave the way for further breeding and hybridization, unlocking the potential for developing novel strains with desirable traits. These discoveries drive innovation in the cannabis industry, catering to evolving consumer preferences and market trends.

Unveiling Autoflowering Markers: A Tale of Two Traits

Autoflower 1: Through meticulous research, the locus responsible for the autoflower trait, along with a significant flowering time locus (Early1), was mapped using bulked segregant analysis. This groundbreaking study led to the development of high-throughput molecular marker assays, shedding light on the genetic underpinnings of photoperiod-insensitive flowering in high-cannabinoid C. sativa accessions.

Autoflower 2: In a recent breakthrough, researchers identified another gene (Autoflower2) linked to autoflowering cannabis through advanced techniques such as whole genome sequencing (WGS) and bulked segregant analysis (BSA). This dominant trait, distinct from Autoflower1, underscores the diverse mechanisms driving autoflowering cannabis evolution. The availability of PACE genetic markers for both approaches heralds a new era in autoflower breeding, offering breeders powerful tools to accelerate their efforts and expand their genetic repertoire.

The journey of autoflowering cannabis epitomizes the spirit of innovation and exploration within the cannabis community. By harnessing the power of genetics and embracing technological advancements, breeders and researchers continue to push the boundaries of what’s possible, shaping the future of cannabis cultivation and enriching the lives of consumers worldwide.

Cannabis sativa definition

Unveiling the Genetic Landscape of Cannabis sativa: A Path to Precision Breeding

Cannabis sativa L., once shrouded by prohibition, now stands as a versatile and promising plant, thanks to recent legalization initiatives that have unlocked avenues for medical research and industrial growth. However, despite its rich history and diverse applications, the cannabis research community has long lagged behind in understanding its genetics and trait inheritance compared to other major crops.

To bridge this gap, a recent groundbreaking genome-wide association study (GWAS) was undertaken, focusing on nine key agronomic and morphological traits. Leveraging a panel of 176 drug-type Cannabis accessions sourced from the Canadian legal market, this study utilized high-density genotyping-by-sequencing (HD-GBS) to generate dense genotyping data. The result? A comprehensive catalogue of 800 K genetic variants, with 282 K common variants earmarked for GWAS analysis.

Through meticulous GWAS analysis, 18 markers significantly associated with agronomic and morphological traits were identified. Some of these markers wielded substantial phenotypic impact, leading researchers to putative candidate genes residing in high linkage disequilibrium (LD) with the markers. These findings lay a robust foundation for innovative cannabis research, poised to leverage genetic markers in informing breeding programs geared towards meeting the diverse needs of the industry.

Genetic insights into cannabis sativa

Advancements in Cannabis Breeding: A Promising Frontier

Modern breeding technologies, akin to those employed in other high-value crops, hold promise in enhancing Cannabis traits to meet evolving needs across manufacturing, medicinal, recreational, and culinary domains. The advent of next-generation sequencing technology (NGS), coupled with powerful bioinformatic tools, has paved the way for genotype-phenotype association studies, facilitating investigations into the relationship between genetic variation and phenotypic traits.

While classic quantitative trait loci (QTL) mapping studies have shed light on maturity-related QTL in both hemp and drug-type Cannabis, modern GWAS analyses offer a more comprehensive approach by identifying loci related to phenotypes within large populations of unrelated individuals. These studies leverage the information of linkage disequilibrium (LD) between QTL and neighbouring genetic markers to pinpoint regions on the genome that influence traits.

Empowering Precision Breeding with Molecular Tools

To empower Cannabis breeders and researchers, this study provides high-value markers linked to essential agronomic and morphological traits, identified through GWAS conducted on 176 drug-type Cannabis accessions. Utilizing the Bayesian-information and linkage disequilibrium iteratively nested keyway (BLINK) statistical method, markers associated with crucial traits were identified, laying the groundwork for a comprehensive understanding of Cannabis genetics.

In conclusion, this pioneering exploration of the genetic landscape of Canadian drug-type Cannabis through GWAS analysis, coupled with high-throughput genotyping and precise phenotyping, opens new vistas for advancing breeding programs and addressing the diverse needs of emerging industries. The identified markers promise to expedite breeding efforts, enabling the cultivation of Cannabis varieties optimized for various purposes and applications, thereby propelling the industry forward into a new era of precision agriculture.

Cannabis sativa breeding
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MEET OUR TEAM

Steve Asquith Managing Director
Steve began his career in the Genetics Division of GlaxoSmithKline, as part of the team establishing GSK’s high-throughput core genotyping laboratory. Steve joined KBioscience when it was first founded in 2002 and was a key driver in taking the company from a small start-up to a multi-national service laboratory, quickly growing the company’s revenue to over $7.5M p.a. Following the acquisition of Kbioscience by LGC in 2011, Steve was appointed Global Director of Operations for LGC Genomics, responsible for over 100 staff in Europe and N. America, successfully elevating the genotyping products and service business. Steve held a crucial leadership role until he left in 2016. In 2017 Steve joined forces with John Holme to create 3CR Bioscience, a new company with a mission to deliver outstanding, customer-focused genotyping products with innovation and affordability at its core.
Dr. John Holme Technical Director
John joined KBioscience shortly after it was founded, in 2003, and became Head of Technical Development, building the company’s genotyping and DNA extraction product portfolio and service delivery until 2011 when it was acquired by LGC. Post-acquisition, John was appointed Head of Technical Group for LGC Genomics, in charge of all Research & Development and Technical Support activities for the company. In this role John continued to build on the high-quality products and services provided to the companies growing customer base. During the 19 years John has worked in commercial R&D, he has co-invented numerous highly successful products including PACE®, ProbeSure, KASP™, KlearKall, KlearGene, KlearAmp and KlearTaq™, creating breakthrough offerings in genotyping and extraction and generating huge revenues for the companies he has worked in. In 2017, he joined forces with Steve Asquith and started 3CR Bioscience. John is dedicated to developing outstanding, innovative genotyping products and providing the very best technical support to customers globally.
Dr. Nisha JainOperations Director
Nisha has been innovating since the start of her career at Geneform Technologies developing Iso-thermal Genotyping Technologies. Nisha joined KBioscience in 2008, as Senior R&D Scientist and key account Technical Support Scientist, developing KASP and Klearkall performance and coinventing two further versions of KASP. Nisha has more than 15 years’ experience working in molecular biology and genotyping technologies, with extensive experience in the areas of R&D, Quality Assurance and Customer Technical Support. She has technically assisted many giants of the industry with their protocol development and troubleshooting and continues to deliver high-quality support and guidance. In 2018, Nisha joined 3CR Bioscience as Operations Director where she continues to develop PACE and ProbeSure for an increasing range of applications, and to grow 3CR Bioscience’s new product pipeline. Nisha is dedicated to developing outstanding, innovative genotyping products and providing the very best technical support to customers globally.
Nazma Saffin General Manager
For 20 years Nazma Saffin has worked and gained extensive expertise within the genotyping sector. Working at Kbioscience and then LGC, she has held operational leadership posts responsible for manufacturing and laboratory services. With experience of ISO 9001 implementation, production scale up and LEAN operations, Nazma has successfully led highly profitable production departments. Joining 3CR Bioscience in 2022, Nazma is committed to delivering operational excellence.
Jon Curtis Non-Executive Chair
After 8 years in The Royal Air Force, Jon moved to the Imperial Cancer Research Fund where he pioneered the use of ultra high-throughput genomic automation, capable of 46,000 PCRs per hour. In the 1990’s Jon joined GlaxoSmithKline, implementing a high-throughput genomics platform into their drug discovery pipeline. Whilst there he also developed acoustic mixing into compound management, becoming the gold standard across pharma. Jon developed the world’s first commercially viable 1536-well PCR plates, automated thermal & laser plate-sealer, plus automated liquid-handling & tip washing tools to reduce waste and costs. In 2002 Jon co-founded KBioscience with Phil Robinson, utilising ultra high-throughput PCR instrumentation & a suite of automation tools to create the company’s SNPline robotic platform, with a capacity of 250,000 PCRs/day. The business was underpinned by their ground-breaking patented genotyping chemistry, KASP™, which has over 10,000 scientific papers to date. In November 2022 Jon joined 3CR Bioscience acting as an advisor bringing his commercial and scientific experience to the company.