Generation of a New Glutinous Photothermosensitive Genic-Male-Sterile (PTGMS) Line by CRISPR/Cas9-Directed Mutagenesis of Wx in Rice (Oryza sativa L.)

Rongbai Li,Yaoguang Liu,Xinying Guo,Xin Wang,Kaichong Teng

doi:10.3390/agriculture11111044

Rongbai Li, Yaoguang Liu + Show 3 more

Open Access

https://doi.org/10.3390/agriculture11111044

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Abstract

The Photothermosensitive Genic-Male-Sterile (PTGMS) line, Y58S, an indica rice variety, combines high-quality and high-light-efficiency use, disease and stress resistance, and excellent plant type and mating force. Y58S is widely used to assemble two-line hybrid rice varieties, especially super hybrids. The Wx gene is the main effector gene for controlling amylose synthesis, which determines the amylose content (AC) of rice grains. By editing this gene, a glutinous line with a low AC can be obtained. In this study, the CRISPR/Cas9 system was used to mediate the editing of the Wx gene, which caused ultra-low AC mutations that produced a PTGMS glutinous rice strain with excellent waxiness. The results showed that 18 positively transformed plants were obtained from the T0 generation, with a mutation rate of 64.29%, of which six were homozygous mutant plants, indicating that the gene-editing target had a higher targeting efficiency and a higher homozygosity mutation rate. Compared to the wild type, the AC of the mutants was significantly lower. Through molecular marker detection and screening of T1 and T2 generations, five homozygous T-DNA-free mutant strains were identified that were consistent with Y58S in fertility and other agronomic traits except for AC. Among these, the AC of the W-1-B-5 homozygous mutant, the glutinous PTGMS line wx-Y58S, was as low as 0.6%. Our research revealed that the Wx gene of excellent PTGMS rice can be edited to generate a new waxy PTGMS line using the CRISPR/Cas9 system. This study provided a simple and effective strategy for breeding high-yield, high-quality, and glutinous two-line hybrid rice, and provided excellent sterile lines for their large-scale application. Once put into use, waxy hybrid rice will greatly improve the yield of glutinous rice and increase social benefits.

Highlights

Introduction iationsMore than half of the world’s population depends on rice as a dietary staple, in China
The results indicate that the mutations in waxy gene (Wx) genes do not change other agronomic traits
The results showed that the pollen-iodine staining of wx-Y58S and Y58S plants that were grown in the fieldinwas that is, theythat wereis,round whilesterile; the pollen of normally grown thereddish-brown, field was reddish-brown, they sterile; were round while wx-Y58S and Y58S plants that were normally grown in cold water Iodine staining was all the pollen of wx-Y58S and Y58S plants that were normally grown in cold water Iodine black-blue, meaning fertility was transformed into fertile (Figure 7A–D)

Summary

Introduction

More than half of the world’s population depends on rice as a dietary staple, in China. It is no longer associated just with satiety, but rather with a better quality of life. The molecular markers of 50 indica-japonica comparisons distributed throughout the. 50 indica-japonica comparisons distributed throughout that 15 weremarkers consistent the Japonica rice variety Nipponbare. Itwere was fact that no markers were detected on chromosomes 1 and 8, other chromosomes found that markers were consistent with the Japonica rice variety Nipponbare. Except detected, and they were mainly concentrated on chromosomes 4 and 5. The remaining for the fact that markers with wereindica detected chromosomes

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