This article has been reviewed according to Science X's editorial process and policies. Editors have highlighted the following attributes while ensuring the content's credibility: A new gene-editing system could make it easier for researchers to introduce precise genetic changes into crops such as tomatoes, soybeans and rapeseed. Developed by researchers in China, SmartPE improves prime editing efficiency in several dicot plants, addressing a major limitation of the technique.
The new system introduces mismatches into the reverse transcriptase templates (RTTs) of two prime editing guide RNAs (pegRNAs), combining this strategy with a cis-regulatory element and an engineered reverse transcriptase. The study, led by professor Gao Caixia from the Institute of Genetics and Developmental Biology (IGDB) of the Chinese Academy of Sciences (CAS), was published in Nature Plants on Oct. 28. Prime editing can install base substitutions and small insertions or deletions without double-strand breaks or a donor template, with relatively few off-target events and byproducts.
In dicot plants, however, its efficiency is low and varies widely among target sites. In this study, the researchers first inserted cis-regulatory elements between the promoter and the Cas9 coding sequence to increase editor expression. The START element performed best, raising transcript levels 2.1-fold and editing efficiency at four Arabidopsis targets by about 1.8-fold.
Building on this improvement, the team tested a dual-pegRNA strategy, but editing efficiency did not improve. The researchers reasoned that complementary pairing between the RTT and genomic DNA may hinder reverse transcription and that mismatch repair (MMR) suppresses prime editing. They therefore introduced complementary mismatched bases into the RTTs of both pegRNAs.
This modification increased average editing activity at the four targets more than sixfold, reaching approximately 10%. The resulting system was named SmartPE. The team further examined how the mismatches should be arranged.
Continuous mismatches or those spaced 3 bp apart yielded the highest editing efficiency (up to 20%), whereas intervals of 6 bp or more sharply reduced efficiency. This pattern differed from that observed in single-pegRNA systems. Mismatches at both ends of the editing window improved efficiency, whereas those in the middle did not, suggesting that mismatch placement may influence the initiation of reverse transcription.
Extract — continue reading at the source.
