Abstract
Key messageCRISPR-Cas9-based genome editing and EMS mutagenesis revealed inter-cultivar differences and additivity in the contribution of TaGW2 homoeologues to grain size and weight in wheat.The TaGW2 gene homoeologues have been reported to be negative regulators of grain size (GS) and thousand grain weight (TGW) in wheat. However, the contribution of each homoeologue to trait variation among different wheat cultivars is not well documented. We used the CRISPR-Cas9 system and TILLING to mutagenize each homoeologous gene copy in cultivars Bobwhite and Paragon, respectively. Plants carrying single-copy nonsense mutations in different genomes showed different levels of GS/TGW increase, with TGW increasing by an average of 5.5% (edited lines) and 5.3% (TILLING mutants). In any combination, the double homoeologue mutants showed higher phenotypic effects than the respective single-genome mutants. The double mutants had on average 12.1% (edited) and 10.5% (TILLING) higher TGW with respect to wild-type lines. The highest increase in GS and TGW was shown for triple mutants of both cultivars, with increases in 16.3% (edited) and 20.7% (TILLING) in TGW. The additive effects of the TaGW2 homoeologues were also demonstrated by the negative correlation between the functional gene copy number and GS/TGW in Bobwhite mutants and an F2 population. The highest single-genome increases in GS and TGW in Paragon and Bobwhite were obtained by mutations in the B and D genomes, respectively. These inter-cultivar differences in the phenotypic effects between the TaGW2 gene homoeologues coincide with inter-cultivar differences in the homoeologue expression levels. These results indicate that GS/TGW variation in wheat can be modulated by the dosage of homoeologous genes with inter-cultivar differences in the magnitude of the individual homoeologue effects.
Highlights
Wheat is one of the most widely grown crops in the world and contributes 20% of the calories in human diets (FAO: http://www.fao.org/faostat/)
We previously showed that the KO of all three homoeologues of TaGW2 gene by CRISPR-Cas9 increased grain size (GS) and thousand grain weight (TGW) much higher than the reported increase in TaGW2A1 mutants (TGW; 28% compared to 7%), which suggests that the B and D homoeologues (TaGW2-B1 and TaGW2D1) have potential phenotypic effects (Wang et al 2018)
Using the wheat TILLING populations and the CRISPRCas9-based genome editing technology, we showed that the KO of each of the three homoeologues of the TaGW2 gene, which are all functional in Bobwhite (Zhang et al 2018), and the A and B genome copies of the TaGW2 gene in Paragon increase GS and TGW
Summary
Wheat is one of the most widely grown crops in the world and contributes 20% of the calories in human diets (FAO: http://www.fao.org/faostat/). The allohexaploid genome of common wheat (Triticum aestivum, 2n = 6x, genomes AABBDD) has many genes present in three copies on homoeologous chromosomes. This genetic redundancy impedes the functional studies of genes controlling important agronomic traits, because the phenotypic effects of loss-of-function mutations in a single homoeologue are frequently masked by other gene copies (Borrill et al 2015). The development of reverse genetics methods based on either TLLING (Krasileva et al 2017) or CRISPR-Cas gene editing (Wang et al 2016, 2018) provide powerful tools to understand the role of wheat gene homoeologues in controlling complex phenotypic traits and to expand genetic diversity accessible for wheat improvement offsetting the negative effects of domestication and improvement bottlenecks on genetic variation (Avni et al 2017; Cavanagh et al 2013; Wang et al 2014a)
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