Base editing in plants: Current status and challenges

Genome editing technologies have revolutionized the field of plant science by enabling targeted modification of plant genomes and are emerging as powerful tools for both plant gene functional analyses and crop improvement. Although homology-directed repair(HDR)is a feasible approach to achieve precise gene replacement and base substitution in some plant species, the dominance of the non-homologous end joining pathway and low efficiency of HDR in plant cells have limited its application. Base editing has emerged as an alternative tool to HDR-mediated replacement, facilitating precise editing of plant genome by converting one single base to another in a programmable manner without a doublestranded break and a donor repair template. In this review, we summarize the latest developments in base-editing technologies as well as their underlying mechanisms. We review current applications of these technologies in plant species. Finally, we address the challenges and future perspectives of this emerging technology in plants.

Highly efficient CRISPR-SaKKH tools for plant multiplex cytosine base editing

Base editing, as an expanded clustered regularly interspaced short palindromic repeats(CRISPR)-Cas genome editing strategy, permits precise and irreversible nucleotide conversion. SaKKH, an efficient variant of a Cas9 ortholog from Staphylococcus aureus(SaCas9), is important in genome editing because it can edit sites with HHHAAT protospacer adjacent motif(PAM) that the canonical Streptococcus pyogenes Cas9(SpCas9) or its variants(e.g. xCas9, Cas9-NG) cannot. However, several technical parameters of SaKKH involved base editors have not been well defined and this uncertainty limits their application. We developed an effective multiplex cytosine base editor(SaKKHn-pBE) and showed that it recognized NNARRT, NNCRRT, NNGRGT, and NNTRGT PAMs. Based on 27 targets tested, we defined technical parameters of SaKKHn-pBE including the editing window, the preferred sequence context, and the mutation type. The editing efficiency was further improved by modification of the SaKKH sgRNA. These advances can be applied in future research and molecular breeding in rice and other plants.

A Clean Technology for Future Prospective: Emission Modeling of Gas Based Power Plant

The aim of present research is to study the dispersion of air pollutants using the air quality model, AERMOD and to predict the impact of pollutants (PM10, NO2 and CO) at the receptor level released from Gas Based Power Plant (GBPP). The net-concentrations including monitored data plus predicted values of PM10, NO2 and CO would be increased from base value 75 to 77.61 μg/m3 with an increase of 3.48%, 22 to 26.66 μg/m3 with an increase of 21.18% and 428 to 538.37 μg/m3 with an increase of 25.79% respectively. The study of hill effect showed that it had profound impact upon the dispersion of pollutants and the ratio (with hill and without hill) of each pollutant was 3.89 for PM10 (24 hr), 2.40 for NO2 (24 hr) and 13.98 for CO (1 hr). The natural gas based plant not only decreases the pollution level but also reduces the hospital treatment cost and protects the public health. The modeling results suggest that the GBPP could be a clean technology as replacement of coal power plants located in the city which pollute the environment considerably in spite of control measures installed.

Evaluation and Analysis for Survey of Tea Production Quality Safety Management and Control

[Objectives]This study was conducted to investigate and evaluate the quality safety of tea production in China.[Methods]With the production of famous and high-quality green tea in the western Sichuan tea areas as the investigation object,the practice survey and comprehensive evaluation analysis method were adopted to conduct research on tea production quality safety management and control,and a comprehensive evaluation was carried out in combination with the survey.[Results]Statistical analysis and scoring were carried out according to relevant indices,and the current situation of tea production quality safety was comprehensively evaluated.Four hazards were found,including pesticide residues,heavy metal residues,harmful microorganisms and non-tea impurities,and improvement measures were proposed.[Conclusions]This study provides a theoretical reference for improving the quality safety of tea production in China.

Comparison of a plant based natural surfactant with SDS for washing of As(V) from Fe rich soil

This study explores the possible application of a biodegradable plant based surfactant, obtained from Sapindus mukorossi, for washing low levels of arsenic （As） from an iron （Fe） rich soil. Natural association of As（Ⅴ） with Fe（Ⅲ） makes the process difficult. Soapnut solution was compared to anionic surfactant sodium dodecyl sulfate （SDS） in down-flow and a newly introduced suction mode for soil column washing. It was observed that soapnut attained up to 86% efficiency with respect to SDS in removing As. Full factorial design of experiment revealed a very good fit of data. The suction mode generated up to 83 kPa pressure inside column whilst down-flow mode generated a much higher pressure of 214 kPa, thus making the suction mode more efficient. Micellar solubilisation was found to be responsible for As desorption from the soil and it followed 1st order kinetics. Desorption rate coefficient of suction mode was found to be in the range of 0.005 to 0.01, much higher than down-flow mode values. Analysis of the PT-IR data suggested that the soapnut solution did not interact chemically with As, offering an option for reusing the surfactant. Soapnut can be considered as a soil washing agent for removing As even from soil with high Fe content.

Single-nucleotide editing for zebra3 and wsl5 phenotypes in rice using CRISPR/Cas9-mediated adenine base editors

The CRISPR/Cas9-mediated base editing technology can efficiently generate point mutations in the genome without introducing a double-strand break(DSB)or supplying a DNA donor template for homology-directed repair(HDR).In this study,adenine base editors(ABEs)were used for rapid generation of precise point mutations in two distinct genes,OsWSL5,and OsZEBRA3(Z3),in both rice protoplasts and regenerated plants.The precisely engineered point mutations were stably inherited to subsequent generations.These single nucleotide alterations resulted in single amino acid changes and associated wsl5 and z3 phenotypes as evidenced by white stripe leaf and light green/dark green leaf pattern,respectively.Through selfing and genetic segregation,transgene-free,base edited wsl5 and z3 mutants were obtained in a short period of time.We noticed a novel mutation(V540A)in Z3 locus could also mimic the phenotype of Z3 mutation(S542P).Furthermore,we observed unexpected non-A/G or T/C mutations in the ABE editing window in a few of the edited plants.The ABE vectors and the method from this study could be used to simultaneously generate point mutations in multiple target genes in a single transformation and serve as a useful base editing tool for crop improvement as well as basic studies in plant biology.