Production of grains with ultra-low heavy metal accumulation by pyramiding novel Alleles of OsNramp5 and OsLsi2 in two-line hybrid rice.

Ensuring rice yield and grain safety quality are vital for human health. In this study, we developed two-line hybrid rice (TLHR) with ultra-low grain cadmium (Cd) and arsenic (As) accumulation by pyramiding novel alleles of OsNramp5 and OsLsi2. We first generated low Cd accumulation restorer (R) lines by editing OsNramp5, OsLCD, and OsLCT in japonica and indica. After confirming that OsNramp5 was most efficient in reducing Cd, we edited this gene in C815S, a genic male sterile line (GMSL), and screened it for alleles with low Cd accumulation. Next, we generated R and GMSL lines with low As accumulation by editing OsLsi2 in a series of YK17 and C815S lines. When cultivated in soils that were heavily polluted with Cd and As, the edited R, GMSL, and TLHR plants showed significantly reduced heavy metal accumulation, while maintaining a relatively stable yield potential. This study provides an effective scheme for the safe production of grains in As- and/or Cd-polluted paddy fields.

Production of aromatic three-line hybrid rice using novel alleles of BADH2.

Aroma is a key grain quality trait that directly influences the market price of rice globally. Loss of function of betaine aldehyde dehydrogenase 2 (OsBADH2) affects the biosynthesis of 2-acetyl-1-pyrroline (2-AP), which is responsible for aroma in fragrant rice. The current study was aimed at creating new alleles of BADH2 using CRISPR/Cas9 gene editing technology under the genetic background of the japonica Ningjing 1 (NJ1) and indica Huang Huazhan (HHZ) varieties. Sensory evaluation and analysis using headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) showed that the grains of the four homozygous T1 lines with new alleles of BADH2 (nj1-cr BADH2 -1, nj1-cr BADH2 -2, hhz-cr BADH2 -1 and hhz-cr BADH2 -2) produced moderate fragrance and had significantly increased 2-AP content compared with wild-types. Moreover, there were no significant differences in the amylose content and gelatinization temperature among the four lines with new alleles of BADH2 to the wild-types. Thereafter, we crossed the HHZ background new alleles of BADH2 with CMS line Taonong 1A (TN1A) to produce a three-line hybrid variety B-Tao-You-Xiangzhan (BTYXZ) with increased grain aroma. The 2-AP content in grains of the improved BTYXZ-1 and BTYXZ-2 reached at 26.16 and 18.74 µg/kg, and the gel consistency of BTYXZ-1 and BTYXZ-2 increased significantly by 9.1% and 6.5%, respectively, compared with the wild-type Tao-You-Xiangzhan (TYXZ). However, the γ-aminobutyric acid (GABA) content in the improved three-line hybrid rice BTYXZ-1 (5.6 mg/100 g) and BTYXZ-2 (10.7 mg/100 g) was significantly lower than that of the TYXZ. These results demonstrated that CRISPR/Cas9 gene editing technology could be successfully utilized in improving aroma in non-fragrant japonica and indica varieties. In addition, the newly developed BADH2 alleles provided important genetic resources for grain aroma improvement in three-line hybrid rice.

Effects of Grain Shape Genes Editing on Appearance Quality of Erect-Panicle Geng/Japonica Rice.

Long grain geng/japonica rice has a higher market preference due its excellent appearance quality. The dense and erect-panicle 1 (dep1) gene has been widely used in the breeding of high-yielding geng/japonica rice cultivars in China. However, this gene causes short and round grain shape thus making it less attractive in global rice markets. Therefore, breeding of high-yielding long-grain geng/japonica rice cultivars by incorporating dep1 with major-effect grain shape gene is of high priority in rice industry. Up to now, multiple grain shape genes' effect mechanism has been clearly elaborated, however, under the background of erect-panicle geng/japonica rice, the effect of major grain shape genes on the appearance quality need to be further clarified, as detailed reports are limited. Utilizing CRISPR/Cas9 technology, a series of near-isogenic lines (NILs) (YF47dep1-gw8, YF47dep1-gs3, YF47dep1-gl7, YF47dep1-qgl3 and YF47dep1-tgw6) in Yanfeng 47(YF47dep1) background were created. Grain appearance and yield components analysis showed that: (1) All NILs' grain length to width ratio was significantly increased compared to that of YF47dep1, excepted YF47dep1-gs3, (2) The chalkiness degree was significantly reduced in all of the NILs, (3) In all of the NILs, YF47dep1-gw8 grains exhibited the greatest length to width ratio and the lowest chalkiness degree, (4) The composition of glume cells and filling characteristics of the endosperm were two key factors contributing grain shape and grain chalk variations, respectively, and v) Owning to a substantial increase in the thousand grain weight, the yields of YF47dep1-gs3 and YF47dep1-tgw6 were significantly higher than that of YF47dep1, whereas YF47dep1-qgl3 exhibited the lowest yield because of a dramatic decrease in the effective panicle number and thousand grain weight. All the results revealed that pyramiding dep1 with major-effect grain shape alleles was an effective approach to improving the appearance quality of erect-panicle geng/japonica rice, owning to both of the appearance quality and yield improvement, GS3 and TGW6 alleles can be applied directly for breeding long-grain shape geng/japonica rice, and editing GW8 resulted in excellent appearance quality but low yield, therefore, this gene would be difficult to use directly but can be considered as the core germplasm resource.

Inorganic arsenic toxicity and alleviation strategies in rice.

Direct or indirect exposure to inorganic arsenic (iAs) in the forms of AsIII (arsenite) and AsV (arsenate) through consumption of As-contaminated food materials and drinking water leads to arsenic poisoning. Rice (Oryza sativa L.) plant potentially accumulates a high amount of iAs from paddy fields than any other cereal crops. This makes it to be a major source of iAs especially among the population that uses it as their dominant source of diet. The accumulation of As in human bodies poses a serious global health risk to the human population. Various conventional methods have been applied to reduce the arsenic accumulation in rice plant. However, the success rate of these techniques is low. Therefore, the development of efficient and effective methods aimed at lowering iAs toxicity is a very crucial public concern. With the current advancement in technology, new strategies aimed at addressing this concern are being developed and utilized in various parts of the world. In this review, we discuss the recent advances in the management of iAs in rice plants emphasizing the use of nanotechnology and biotechnology approaches. Also, the prospects and challenges facing these approaches are described.

CRISPR-Based Crop Improvements: A Way Forward to Achieve Zero Hunger.

Zero hunger is one of the sustainable development goals set by the United Nations in 2015 to achieve global food security by 2030. The current harvest of crops is insufficient; feeding the world's population and meeting the goal of zero hunger by 2030 will require larger and more consistent crop production. Clustered regularly interspaced short palindromic repeats-associated protein (CRISPR-Cas) technology is widely used for the plant genome editing. In this review, we consider this technology as a potential tool for achieving zero hunger. We provide a comprehensive overview of CRISPR-Cas technology and its most important applications for food crops' improvement. We also conferred current and potential technological breakthroughs that will help in breeding future crops to end global hunger. The regulatory aspects of deploying this technology in commercial sectors, bioethics, and the production of transgene-free plants are also discussed. We hope that the CRISPR-Cas system will accelerate the breeding of improved crop cultivars compared with conventional breeding and pave the way toward the zero hunger goal.