Prime Editing Optimized For Rice And Wheat

Prime editing, a new genome editing approach, has great potential for plant breeding and functional genomic research.

AsianScientist (Apr. 21, 2020) – A research team in China has successfully modified prime gene editing so that it can be used on commercially important plants like rice and wheat. Their findings have been published in Nature Biotechnology.

Over the course of hundreds of thousands of years of domestication, most of the plants we eat today have become vastly different from their wild counterparts. Being able to precisely manipulate the genetic material of these plants could render them resistant to drought or pests, or even make them more nutritious.

Existing techniques such as nuclease-initiated homology-director repair (HDR) can generate useful point mutations and insertions/deletions (indels), but are currently inefficient. In human cells, prime editing, which uses engineered Cas9 nickase (H840A)-reverse transcriptase (RT) fusion proteins paired with a prime editing guide RNA (pegRNA), has shown promise as an efficient and accurate gene editing tool. However, prime editing has yet to be applied to plants.

In the present study, a research team led by Professor Guo Caixia of the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences reported the optimization of a prime editing system for creating desired point mutations, insertions and deletions in two major cereal crops, namely, rice and wheat. The main components of the system are a Cas9 nickase-RT fusion protein and a pegRNA.

Using their prime editing system, these researchers produced all 12 kinds of single base substitutions, as well as multiple point mutations and small DNA insertions and deletions at nine rice and seven wheat sites in protoplasts, with efficiencies up to 19.2 percent. The efficiency of the editing was strongly affected by the length of the primer binding site and RT template.

Although their prime editing approach introduced off-target effects, these could be reduced by optimizing RT template length. Moreover, using a prime editing system optimized for plants, the researchers found that the original RT could be replaced by CaMV-RT (from the cauliflower mosaic virus) and retron-derived RT (from E. coli BL21).

Furthermore, Gao and her collaborators were able to create stable rice plants with mutations that are very difficult to produce with existing gene editing tools, sucha s G-to-T point mutations and multinucleotide substitutions.

“Although the efficiency of the prime editing system is lower than that of base editors, it is still an appealing new tool for creating all 12 types of single-point mutation, mixtures of different substitutions as well as insertions and deletions. The system thus has great potential for plant breeding and functional genomics research,” Gao concluded.

The article can be found at: Lin et al. (2020) Prime Genome Editing in Rice and Wheat.


Source: Chinese Academy of Sciences; Photo: Pexels.
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