Plant biologists have developed a method for growing transgenic and gene-edited plants that cuts the slow and expensive process down from months to weeks. Publishing November 6 in the Cell Press journal Molecular Plant, the method takes advantage of plants' natural ability to regenerate after being wounded or pruned. By injecting bacteria carrying genetic instructions for wound healing and regeneration into a pruned plant's wound site, the researchers triggered the plant to grow new shoots, some of which were transgenic and gene edited. The method shows potential even in species that are usually difficult or impossible to regenerate, such as soybeans.
"Plant regeneration has long been a major limitation in crop biotechnology," says senior author and plant genomicist Gunvant Patil of Texas Tech University. "Our method leverages the plant's inherent regenerative capacity to rapidly produce gene-edited shoots, bypassing months of traditional tissue culture. This innovation has the potential to redefine how we create next-generation, improved crop varieties."
Typically, when creating transgenic or gene-edited plants, biologists edit the DNA of an individual plant cell, which they then stimulate to grow into a new plant. For species like tomatoes that are relatively easy to regenerate, this process takes at least four months; for more difficult species like cotton, it can take almost a year; and for some plants, such as beans and peppers, tissue culture is extremely difficult.
To speed up this process, the researchers took advantage of plants' natural ability to regenerate after being damaged or pruned. When a plant gets wounded, it triggers a molecular cascade that first seals the wound by creating a hard callus and then stimulates cell division and differentiation to replace the lost tissue. This process is regulated by a protein called WIND1, which activates a string of other proteins that are involved in cell differentiation and shoot growth.
"It's like a relay: once WIND1 is activated, it will activate the next step, ESR1, which then activates the next step," says Patil.
To simultaneously induce regeneration and introduce new DNA into plants, the researchers used a plant-infecting bacteria, Agrobacterium, that naturally introduces its own DNA into plant cells. They inserted the genetic instructions for WIND1, ESR1, and several other regeneration genes into Agrobacterium, along with a gene that causes red coloration. Then, they pruned plants and applied the genetically modified Agrobacterium to the wound site.
"You decapitate the plant, you inoculate with Agrobacterium, and then the shoots that grow out of the wound will give rise to seeds that are transgenic or gene edited," says coauthor and plant genomicist Luis Herrera-Estrella of Texas Tech University. "This technique could help us transform species that are usually very difficult to grow in tissue culture because it's faster and more natural."
The researchers first tested the technique in tobacco plants, which are an easy species to regenerate. They showed that applying Agrobacterium accelerated callus formation and the growth of new shoots and successfully produced red transgenic shoots around 35% of the time. When they tested the method in tomatoes, which are slightly more difficult to grow, it produced transgenic plants 21% of the time.
The method was initially unsuccessful in soybeans, which are notoriously difficult to regenerate, but the researchers were able to make it work with some tweaks. Instead of applying Agrobacterium to pruned shoots, they applied the bacteria to soybean seeds that had been stimulated to germinate. Then, they grew the soybeans in the tissue culture for 3.5 weeks before transferring them to soil. With this modified method, the researchers were able to produce transgenic shoots 28% of the time.
"With the conventional method, we need to grow soybeans in tissue culture for at least 3 to 4 months, so reducing that time to 3.5 weeks is a huge advancement," says Patil. "This is the first step, and we are now working to fine-tune this technology to apply it to more difficult crops, such as chickpeas, common bean, and many other crops."
