Brazil develops non-transgenic CRISPR edited sugarcane

18-01-2022 | Crop solutions | News

Flex II sugarcane in a Embrapa test field. - Photo: Embrapa

Scientists from Embrapa Agroenergia in Brazil have developed a non-transgenic CRISPR edited sugarcane.The edited sugarcane varieties are called Cana Flex I and Cana Flex II. The respective distinctive features are easier cell wall digestibility and higher sucrose concentration in plant tissues.

The edited plants are considered non-transgenic, or DNA-Free, according to Normative Resolution No. 16 (RN No. 16) of the National Biosafety Technical Commission (CTNBio), issued on 12/9/2021. Both developments used the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats), a “Nobel prized” and revolutionary gene manipulation technique discovered in 2012.

They respond to one of the biggest scientific challenges in the sugar cane sector: easy enzyme’s access into sugars trapped in cells, which facilitates ethanol production (first and second generation), and better extraction of other bioproducts.

In the case of Cana Flex I, the CRISPR technique silenced the gene responsible for the rigidity of the plant cell wall, in order to increase its “digestibility”. This means the enzymatic hydrolysis process – a chemical process that extracts compounds from plant biomass – is more efficient.

Cana Flex II: more sucrose

The second variety, Cana Flex II, was created by silencing a gene in the sugarcane tissue. As a result, the researchers achieved a considerable increase in sucrose production in the plant stems.

“Once we had found that characteristic of sugar accumulation, we transferred this knowledge to the sugarcane crop, which is the target of our research”, explains Embrapa researcher Hugo Molinari. According to him, they observed 15% more sucrose in the sugarcane stalk and a increase of 200% in the leaves. The study also identified a higher concentration of other sugars, such as glucose and fructose, both in juice and plant tissue.

“We also carried out tests to see if the gene had an effect on improving saccharification, which is the conversion of cellulose into industrial sugar. Once again, we observed an increase of around 12%”, Molinari said.

Bioethanol production

According to Molinari the main advantages of Cana Flex II are higher efficiency in bioethanol production, better varieties for industrial purposes, greater digestibility for animal feed and more value added for the sector as a whole.

“In 2020/2021, the world´s total production of sugar was 188 million tonnes, with Brazil being responsible for 39 million tons, equivalent to 21% of the global sugar production”, says Molinari.

He also said sugarcane cultivation can contribute to a cleaner energy matrix. “Today we know more than 45% of the Brazilian energy matrix is renewable and sugarcane contributes 30% to these renewable sources”, Molinari said.

Also read: Future of CRISPR technology in crops reviewed

The world’s first CRISPR edited sugarcane was developed earlier in the USA

Led by Fredy Altpeter, professor of Agronomy at the University of Florida, a group of scientists have used CRISPR to turn off magnesium chelatase genes in sugarcane varieties and produce quickly identifiable plants with light green to yellow leaves. These plants may require less nitrogen with the same crop performance. This study has been published in Frontiers in Genome Editing.
The group also created an efficient and reproducible gene targeting in sugarcane for Crispr usage. This second work demonstrated a precise substitution of multiple copies of the target gene with a superior version, conferring herbicide resistance to the new sugarcane variety. This second study also has been published in Frontiers in Genome Editing.
According to Fredy Altpeter, based on PCR analysis they also identified edited events that did not result in transgene detection from genomic DNA. “But for a statement that they are 100% transgene free one would either have to fully sequence the entire genome which is quite an effort for the complex sugarcane genome (and we have not done yet) or use a gene editing approach that does not involve DNA (I assume that this is the approach that Embrapa took).”
Dr. Altpeter said the regulatory process is different in Brazil and the U.S. “In contrast to Brazil, the U.S. gene edited events without transgene do not require regulatory approval by the US regulatory agencies they are treated as non-gmo (inquiry is optional).
Embrapa states it does not claim to be the first institution to use the genomic editing technology via CRISPR/Cas9 system in sugarcane. “Other groups have used the technology in the crop, with the group led by Prof. Fredy Altpeter being the pioneer,” Embrapa said. According to Embrapa, not every event obtained through genomic editing techniques, among them the CRISPR/Cas9 system, produces non-transgenic plants as a result. “The Brazilian legislation establishes, through Normative Resolution No. 16 of the National Biosafety Technical Commission (CTNBio), that such classification only occurs after case-by-case analysis by CTNBio. Embrapa Agroenergy submitted two genetically edited varieties, Cana Flex I and II, for CTNBio’s appreciation. After analysis, both were considered non-transgenic.“
According to Embrapa, the text available on the Embrapa website states that Cana Flex I and II varieties were given unprecedented approval by the Brazilian regulatory agency for being considered non-transgenic. “Therefore, it informs that the group led by Dr. Hugo Molinari, from Embrapa Agroenergy, was the first to obtain and approve non-transgenic varieties of genetically edited sugarcane. Thus, this is not a disclosure of the “first use of the technique” or of its “proof of concept” in sugarcane.” Embrapa states they “are not aware of any other regulatory agency in the world that has approved any variety of sugarcane obtained by the CRISPR/Cas9 tool as non-transgenic.”