Abstract: Soil salinization has emerged as a major threat affecting crop yields. Global warming leads to a massive loss of terrestrial water and makes soils saltier. Cultivating salt-tolerant crops is the major strategy adopted for utilizing these salinized soils. Sea Rice 86 (SR86) is one such elite salt-tolerant rice variety derived from ancient indica rice. However, SR86 has multiple wild traits, such as tallness and strong photoperiod sensitivity (PS), which have limited its application in agricultural production. In this study, we edited 13 genes responsible for 10 traits in SR86 to develop an improved SR86M line by using clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9 multiplex-genome-editing technology, high-throughput sequencing, crossing, and progeny selection. Subsequent analysis of SR86M detected nine genes with expected mutations, leading to changes in seven traits, including improvements of plant architecture, plant height and PS decreased, grain number, grain length, fragrance, and nitrogen utilization efficiency increased. The improved agronomic traits in SR86M are similar to modern cultivated rice, along with elite salt tolerance like SR86, indicating suitability for potential cultivation. Our results also reveal the efficiency of multiplex-genome-editing in directional improvement of crop varieties.

Key words: CRISPR/Cas9 technology, directional improvement, multiplex‐genome‐editing, salt‐tolerant rice, Sea Rice 86