CRISPR-Cas9 gene editing technology—sometimes billed as “programmable DNA scissors”—is still relatively new in the world of plant breeding. But with its speed, precision, and regulatory advantages in some markets at least (enabling more rapid commercialization vs crops classed as GMOs), it is gaining traction.

AgFunderNews caught up with Dr. Paulo Arruda, professor of genetics at the State University of Campinas in São Paulo, and founder and CEO of startup InEdita Bio, to explore how gene editing can boost crop resilience to drought and pests.

AFN: Tell us about why you started InEdita Bio.

PA: The way we produce food nowadays is not sustainable. More than 80% of the energy the human population gets comes from a handful of crops. And 20% of the grains that are produced are lost to diseases, mostly fungi and pests, although hundreds of millions of tons of pesticides are sprayed to control the problem. We cannot keep doing things the same way.

AFN: What’s the attraction of CRISPR?

PA: CRISPR allows us to use an enzyme, a nuclease that is guided by guide RNAs to a specific point in the genome of any organism and make small changes so that we can change a plant from a susceptibility state to a resistance state.

During the breeding process over the years, we’ve lost the genetic variability that confers resistance to pests and disease because breeding has been focused on [increasing] yield. But now we need technology to help plants grow in more challenging environments.

AFN: Are single genes typically involved in conferring these desired traits or do they involve multiple genes?

PA: Most of the characteristics involved in yield are what we call polygenic where several genes are involved. Each gene adds a small piece, a small percentage of the yield. But when we talk about disease resistance, a single gene can confer resistance to fungi, bacteria and viruses, but the resistance alleles of those genes have been lost in the breeding programs.

Let’s take corn as an example. Here in Brazil there are over 200 commercial hybrids with different genetic backgrounds that are grown from the south to the north of the country. So when you talk about drought resistance, for example, you need to take into account those different areas and regions and different germplasm, there is a complex genetic and environmental combination.

The beauty of gene editing is that we can play with a single gene but we can play also with multiple genes using multiplex genome editing [using multiple guide RNAs to target multiple genomic sites simultaneously] which can include multiple different genes or multiple sites within the same gene or multiple alleles of a gene].

We can also use this technology to add specific modifications on commercial germplasm, varieties and hybrids that are grown in a particular region.

AFN: I understand that InEdita Bio is looking closely at the microbial populations that live with plants?

PA: We use gene editing for developing relevant traits in plants, but plants are also associated with thousands of microorganisms that help them with nutrient uptake and resistance to drought, stress, and diseases.

For example, in Central Brazil, you have areas where you can go six months without any rain, but there’s huge biodiversity. So, the question is how? The answer is that the plants have developed mechanisms to survive, but also, they are associated with microorganisms that live in this area that help them to survive in poor environments in terms of nutrients and water availability.

So, we need to put together plant genetics with microbial genetics. This is the future of biotechnology.

And this is some of what we are doing at InEdita, genome editing to improve the association of the microorganism with the plant. So, you can do genome editing in the microorganism and you can do genome editing in plants and in the future, we’re going to see this merging of plant genetics with microbial genetics. Exploring this idea, I think, will help increase the sustainability of food production.

For example, we’re working on developing plants that better associate with nitrogen-fixing microbes. Soybeans for example can associate with nitrogen-fixing bacteria that produce nodules in the roots, and supply all the nitrogen through biological nitrogen fixation (BNF). In Brazil, the largest producer of soybeans in the world, we don’t need to supply the crop with nitrogen fertilizers because the nitrogen comes from BNF. [Brazil is known for very efficient biological nitrogen fixation in soybeans, thanks to widespread inoculation with effective strains of Bradyrhizobium and the selection of soybean varieties well adapted to symbiosis].

So, can we make other crops [which do not naturally form root nodules or enter into a symbiotic relationship with nitrogen-fixing bacteria] that can attract bacteria in the same way?

AFN: How can you make plants that attract these kinds of beneficial bacteria?

PA: You can make plants that produce signaling compounds that attract the beneficial microorganisms and help them to grow and colonize and help plants to fix nitrogen. This is the future.

AFN: How is InEdita making its edits? You’re not using tissue culture?

PA: You can deliver CRISPR systems using biological vectors such as viruses. You can engineer viruses to deliver guide RNAs and so forth. We have developed a system where you can bypass tissue culture [which is labor-intensive, plus not all plants regenerate well from tissue culture] by delivering CRISPR into cells by particle bombardment.

Particle bombardment is a machine where you can have very small gold particles, and then you can [use them as carriers] for DNA that is going to express a nuclease and guide RNA. And then these particles are shot into plant cells allowing the DNA to enter the cells. And then you can recover seeds [carrying the genetic change].

AFN: Are you bombarding cells in culture or in fully-grown plants, or something else?

PA: You can do this in a growing plant, where you have, for example, meristematic tissues [to induce inheritable edits directly in the meristem, a region of a plant comprising undifferentiated cells from which the rest of the plant—including the germline—develops].

And then you just shoot [the particles] into this meristematic tissue and leave plants to develop shoots from that tissue. Other approaches are putting Arabidopsis flowers [a plant in the mustard family] into a mixture of agrobacteria [a widely used method for agrobacterium-mediated transformation] and then harvest the seeds. But Arabidopsis is not corn, it’s not soybean, it’s not wheat [so this approach may not work for many crops].

AFN: You’re deploying RNAi (RNA interference)?

PA: We have developed a system for delivering RNAi [RNA interference] against diseases impacting plants by [engineering the] non-translated [non-coding] regions of plant genes [DNA sequences that are not translated into proteins but play critical roles in gene regulation, for example].

RNAi is a natural gene-silencing mechanism found in all organisms to fight against disease, which plants can use to defend against pathogens by silencing specific genes in fungi, bacteria, or insects. We enable the plant to produce RNAi molecules that silence key genes in pathogens and enhance resistance.”

AFN: What is your business model?

PA: We develop the technology and we license it to seed companies. We do also co-development projects. For example, now we have two such projects with corn seed companies that are interested in some traits we’re working on that can be used in their elite commercial varieties.

So [in these kinds of projects] there would be some upfront payment to help us develop the project, and then we would have royalties [on an ongoing basis].

AFN: I understand you’re working on Asian Soybean Rust (ASR)?

PA: This is a major problem. We have received grant funding from FINEP, a Brazilian federal funding institution to develop soybean resistant to ASR. This is a project that we want to develop alone [in-house] up to the stage where we have a proof of concept in multiple location field trials, so that we can then negotiate with the major soybean seed companies, not only in Brazil, but elsewhere.

But what is very important is that the technology we’re developing for ASR can also be used to tackle other diseases on other crops. We are using ASR as a demonstration model. This is a major economic problem and a major environmental problem.

AFN: What’s the mechanism of action in your approach to tackling ASR?

PA: We are asking what are the genes that are essential for the pathogen to live? What is the metabolism? If you know what those genes are, you can say, let’s silence those genes. This is what we do.

AFN: Can you speak about the regulatory pathway for gene edited crops?

PA: We are still in the very beginning, although there are some varieties already approved in Brazil for commercialization [a CRISPR-edited soybean from Embrapa with inactivated lectin genes, reducing an anti-nutritional factor in feed, and GDM’s CRISPR-edited drought‑tolerant soybean].

In general, people say [gene edited crops should be exempt from more challenging regulations that apply to transgenic crops] if you are doing what nature could do, whether it’s with a single gene or multiple genes, because nature works with multiple genes, right? We need to decide what we want to do: just keep on spraying to tackle these diseases, or use genetics?

AFN: What’s the long-term potential of gene editing in plant breeding?

PA: It’s going to be transformative. The genome is like a text. There are some places where you can change some words, add a comma. But how many of those modifications do you need to have plants that are more resilient to stress, to disease?

You need to have what I call industrial genetics, and a pipeline where you can, say, correct the alleles that have been mutated or have been selected through breeding programs where people were looking to other traits [increasing yield] and lost some important traits, like, for example, resistance to abiotic stress [drought, heat, salinity etc].

In the past, it has taken, say, 10-15 years to develop a single variety. Imagine a system where we can edit the text of plant genomes to bring all these important traits together, now that we have access to computational biology, AI, and machine learning. We can sequence the genomes of thousands of individuals within wild populations and species that lives in very harsh environments and use the information to ‘correct the text’ of commercial varieties that have been developed by breeders.