Engineering Quantitative Trait Variation for Crop Improvement by Genome Editing

Abstract

Major advances in crop yields are needed in the coming decades. However, plant breeding is currently limited by incremental improvements in quantitative traits that often rely on laborious selection of rare naturally occurring mutations in gene-regulatory regions. Here, we demonstrate that CRISPR/Cas9 genome editing of promoters generates diverse cis-regulatory alleles that provide beneficial quantitative variation for breeding. We devised a simple genetic scheme, which exploits trans-generational heritability of Cas9 activity in heterozygous loss-of-function mutant backgrounds, to rapidly evaluate the phenotypic impact of numerous promoter variants for genes regulating three major productivity traits in tomato: fruit size, inflorescence branching, and plant architecture. Our approach allows immediate selection and fixation of novel alleles in transgene-free plants and fine manipulation of yield components. Beyond a platform to enhance variation for diverse agricultural traits, our findings provide a foundation for dissecting complex relationships between gene-regulatory changes and control of quantitative traits.

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

“…By applying this system, sgRNA arrays for multiplexing applications can also be generated since these ribozyme-based arrays undergo selfcatalyzed cleavage after transcription which, in the naturally occurring system, requires RNaseIII activity [14,15]. The targeting range of Cas9 is not limited to protein coding genes, but noncoding RNAs and regulatory elements can also be targeted [17,18]. Interestingly, for SpCas9 targeted mutagenesis frequencies could also be strongly enhanced via incubation at 37°C and this is applicable to different plant species [16].…”

Genomics-assisted breeding – A revolutionary strategy for crop improvement

“…Genome editing on the gene promoter region could give rise to differential cis-transcription alleles, providing beneficial quantitative variation for plant breeding. This was confirmed by evaluating numerous promoter variants for genes regulating tomato yield traits (Rodríguez-Leal et al 2017). It could be used for the quick isolation of new alleles and finely the control of yield traits.…”

Advancing biotechnology with CRISPR/Cas9: recent applications and patent landscape

“…In the field of crop engineering, CRISPR techniques are currently having a major impact, facilitating cheaper, faster, and more precise engineering in comparison to laborious and time-consuming traditional methods [3,82]. However, it has yet to be determined whether CRISPR based gene editing of crops will be regulated the same way traditional genetically engineered crops are, which ultimately will settle its commercial value within this sector.…”