Efficient genome editing in rice with miniature Cas12f variants

Genome editing,particularly using the CRISPR/Cas system,has revolutionized biological research and crop improvement.Despite the widespread use of CRISPR/Cas9,it faces limitations such as PAM sequence requirements and challenges in delivering its large protein into plant cells.The hypercompact Cas12f,derived from Acidibacillus sulfuroxidans(AsCas12f),stands out due to its small size of only 422 amino acids and its preference for a T-rich motif,presenting advantageous features over SpCas9.However,its editing efficiency is extremely low in plants.Recent studies have generated two AsCas12f variants,AsCas12f-YHAM and AsCas12f-HKRA,demonstrating higher editing efficiencies in mammalian cells,yet their performance in plants remains unexplored.In this study,through a systematic investigation of genome cleavage activity in rice,we unveiled a substantial enhancement in editing efficiency for both AsCas12f variants,particularly for AsCas12f-HKRA,which achieved an editing efficiency of up to 53%.Furthermore,our analysis revealed that AsCas12f predominantly induces deletion in the target DNA,displaying a unique deletion pattern primarily concentrated at positions 12,13,23,and 24,resulting in deletion size mainly of 10 and 11 bp,suggesting significant potential for targeted DNA deletion using AsCas12f.These findings expand the toolbox for efficient genome editing in plants,offering promising prospects for precise genetic modifications in agriculture.

Characterization of grain cadmium concentration in indica hybrid rice

As a consequence of contamination of soil with heavy metals,cadmium accumulation in grain is of great concern worldwide,but especially in southern China.It is important to evaluate the Cd accumulation potential of grain before or when examining and approving new cultivars.An evaluation method and criteria for verifying Cd accumulation potential in rice are proposed,and the Cd accumulation potential of 56 mid-season indica hybrids collected from the provincial cultivar trials in 2016 were investigated.Genotype,environment and their interactions strongly affected the variation in grain Cd accumulation.Two hybrids were identified as slightly Cd accumulating.Hybrids with slight Cd accumulation potential would be suitable for safe grain production on polluted land(total Cd under 2.0 mg$kg–1)in Hunan Province(China)and should be considered for new cultivar evaluation and approval.This evaluation method and criterion could be applied for certifying Cd accumulation potential of rice cultivars.