MIB2 functions in oocyte meiosis by modulating chromatin configuration.

Chromatin configuration serves as a principal indicator of GV (germinal vesicle)-stage oocyte quality. However, the underlying mechanisms governing the chromatin configuration transition from NSN (non-surrounded nucleolus) to SN (surrounded nucleolus) remain unclear. In this study, by conducting a quantitative proteomic analysis, we identified an increased expression of the MIB2 (MIB E3 ubiquitin protein ligase 2) protein in SN oocytes. Specific depletion of MIB2 in SN oocytes not only leads to severe disruption of the meiotic apparatus and a higher incidence of aneuploidy, but also adversely affects meiotic maturation and early embryo development. Notably, overexpression of MIB2 in NSN oocytes facilitates the chromatin configuration transition. Meantime, we observed that forced expression of MIB2 in NSN oocytes significantly mitigates spindle/chromosome disorganization and aneuploidy. In summary, our results suggest that chromatin configuration transition regulated by MIB2 is crucial for oocyte to acquire the developmental competence.

Maternal Obesity Induces the Meiotic Defects and Epigenetic Alterations During Fetal Oocyte Development.

It has been widely reported that obesity adversely impacts reproductive performance of females. However, the effects of maternal obesity on fetal germ cells remain poorly understood. In the present study, by employing a high-fat diet (HFD)-based mouse model, it is discovered that maternal obesity disrupts the chromosomal synapsis and homologous recombination during fetal oogenesis. Moreover, transcriptomic profiling reveales the potential molecular network controlling this process. Of note, the global hypermethylation of genomic DNA in fetal oocytes from obese mouse is detected. Importantly, time-restricted feeding (TRF) of obese mice not only ameliorate the meiotic defects, but also partly restore the epigenetic remodeling in fetal oocytes. In sum, the evidence are provided showing the deficit fetal oogenesis in obese mother, implicating a mechanism underlying the intergenerational effects of environmental insults. TRF may represent a potentially effective approach for mitigating fertility issues in obese patients.

Natural variation in MORE GRAINS 1 regulates grain number and grain weight in rice.

Grain yield is determined mainly by grain number and grain weight. In this study, we identified and characterized MORE GRAINS1 (MOG1), a gene associated with grain number and grain weight in rice (Oryza sativa L.), through map-based cloning. Overexpression of MOG1 increased grain yield by 18.6%-22.3% under field conditions. We determined that MOG1, a bHLH transcription factor, interacts with OsbHLH107 and directly activates the expression of LONELY GUY (LOG), which encodes a cytokinin-activating enzyme and the cell expansion gene EXPANSIN-LIKE1 (EXPLA1), positively regulating grain number per panicle and grain weight. Natural variations in the promoter and coding regions of MOG1 between Hap-LNW and Hap-HNW alleles resulted in changes in MOG1 expression level and transcriptional activation, leading to functional differences. Haplotype analysis revealed that Hap-HNW, which results in a greater number and heavier grains, has undergone strong selection but has been poorly utilized in modern lowland rice breeding. In summary, the MOG1-OsbHLH107 complex activates LOG and EXPLA1 expression to promote cell expansion and division of young panicles through the cytokinin pathway, thereby increasing grain number and grain weight. These findings suggest that Hap-HNW could be used in strategies to breed high-yielding temperate japonica lowland rice.

KAS-seq profiling captures transcription dynamics during oocyte maturation.

In fully grown oocytes, the genome is considered to be globally transcriptionally silenced. However, this conclusion is primarily derived from the results obtained through immunofluorescence staining or inferred from the highly condensed state of chromosomes, lacking more direct evidence. Here, by using a kethoxal-assisted single-stranded DNA sequencing (KAS-seq) approach, we investigated the landscape of single-strand DNA (ssDNA) throughout the genome and provided a readout of the activity and dynamics of transcription during oocyte meiotic maturation. In non-surrounded nucleolus (NSN) oocytes, we observed a robust KAS-seq signal, indicating the high transcriptional activity. In surrounded nucleolus (SN) oocytes, the presence of ssDNA still persists although the KAS-seq signal was relatively weak, suggesting the presence of transcription. Accompanying with the meiotic resumption, the transcriptional activity gradually decreased, and global repression was detected in matured oocytes. Moreover, we preformed the integrative genomics analysis to dissect the transcriptional dynamics during mouse oocyte maturation. In sum, the present study delineates the detailed transcriptional activity during mammalian oocyte maturation.

Multi-omics reveal the metabolic patterns in mouse cumulus cells during oocyte maturation.

Bi-directional communication between cumulus cells and the surrounded oocytes is important for the development and functions of both compartments. However, the metabolic framework in cumulus cells has not been systematically described. In the present study, cumulus cells from cumulus-oocyte complexes (COCs) at three key time points were isolated (arrested GV stage, post-hCG 0h; meiotic resumption GVBD stage, post-hCG 3h; and metaphase II stage, post-hCG 12h), and the temporal metabolomic and proteomic profiling were performed. Integrated multi-omics analysis reveals the global metabolic patterns in cumulus cells during mouse oocyte maturation. In particular, we found the active hyaluronic acid metabolism, steroid hormone synthesis, and prostaglandin E2 (PGE2) production in cumulus cells. Meanwhile, accompanying the oocyte maturation, a progressive increase in nucleotide and amino acid metabolism was detected in the surrounding cumulus cells. In sum, the data serve as a valuable resource for probing metabolism during terminal differentiation of ovarian granulosa cells, and provide the potential biomarkers for improving and predicting oocyte quality.

L-type lectin receptor-like kinase OsCORK1 as an important negative regulator confers copper stress tolerance in rice.

Copper (Cu) is vital for plant growth but becomes toxic in excess, posing potential threats to human health. Although receptor-like kinases (RLKs) have been studied in plant response to abiotic stresses, their roles in Cu stress response remain poorly understood. Therefore, we aimed to evaluate Cu toxicity effects on rice and elucidate its potential molecular mechanisms. Specifically, rice lectin-type RLK OsCORK1 (Copper-response receptor-like kinase 1) function in Cu stress response was investigated. RNA sequencing and expression assays revealed that OsCORK1 is mainly expressed in roots and leaves, and its expression was significantly induced by Cu stress time- and dose-dependently. Kinase activity assays demonstrated OsCORK1 as a Mn2+-preferred functional kinase. Genetically, OsCORK1 gene-edited mutants exhibited increased tolerance to Cu stress and reduced Cu accumulation compared to the wild type (WT). Conversely, OsCORK1 overexpression compromised the Cu stress tolerance observed in OsCORK1 gene-edited mutants. OsCORK1 gene-edited mutants slightly damaged the root tips compared to the WT under Cu stress. Furthermore, OsCORK1 was demonstrated to modulate Cu stress tolerance by mainly altering cell wall components, particularly lignin, in rice. Overall, OsCORK1 is an important negative regulator of Cu stress tolerance, providing a potential gene target to reduce Cu pollution in rice production.

OsMADS5 interacts with OsSPL14/17 to inhibit rice root elongation by restricting cell proliferation of root meristem under ammonium supply.

Nitrogen (N) is a vital major nutrient for rice (Oryza sativa). Rice responds to different applications of N by altering its root morphology, including root elongation. Although ammonium ( NH 4 + ) is the primary source of N for rice, NH 4 + is toxic to rice roots and inhibits root elongation. However, the precise molecular mechanism that NH 4 + -inhibited root elongation of rice is not well understood. Here, we identified a rice T-DNA insert mutant of OsMADS5 with a longer seminal root (SR) under sufficient N conditions. Reverse-transcription quantitative PCR analysis revealed that the expression level of OsMADS5 was increased under NH 4 + compared with NO 3 - supply. Under NH 4 + conditions, knocking out OsMADS5 (cas9) produced a longer SR, phenocopying osmads5, while there was no significant difference in SR length between wild-type and cas9 under NO 3 - supply. Moreover, OsMADS5-overexpression plants displayed the opposite SR phenotype. Further study demonstrated that enhancement of OsMADS5 by NH 4 + supply inhibited rice SR elongation, likely by reducing root meristem activity of root tip, with the involvement of OsCYCB1;1. We also found that OsMADS5 interacted with OsSPL14 and OsSPL17 (OsSPL14/17) to repress their transcriptional activation by attenuating DNA binding ability. Moreover, loss of OsSPL14/17 function in osmads5 eliminated its stimulative effect on SR elongation under NH 4 + conditions, implying OsSPL14/17 may function downstream of OsMADS5 to mediate rice SR elongation under NH 4 + supply. Overall, our results indicate the existence of a novel modulatory pathway in which enhancement of OsMADS5 by NH 4 + supply represses the transcriptional activities of OsSPL14/17 to restrict SR elongation of rice.

BPB1 regulates rice (Oryza sative L.) panicle length and panicle branch development by promoting lignin and inhibiting cellulose accumulation.

Panicle structure is one of the most important agronomic traits directly related to rice yield. This study identified a rice mutant basal primary branch 1 (bpb1), which exhibited a phenotype of reduced panicle length and arrested basal primary branch development. In addition, lignin content was found to be increased while cellulose content was decreased in bpb1 young panicles. Map-based cloning methods characterized the gene BPB1, which encodes a peptide transporter (PTR) family transporter. Phylogenetic tree analysis showed that the BPB1 family is highly conserved in plants, especially the PTR2 domain. It is worth noting that BPB1 is divided into two categories based on monocotyledonous and dicotyledonous plants. Transcriptome analysis showed that BPB1 mutation can promote lignin synthesis and inhibit cellulose synthesis, starch and sucrose metabolism, cell cycle, expression of various plant hormones, and some star genes, thereby inhibiting rice panicle length, resulting in basal primary branch development stagnant phenotypes. In this study, BPB1 provides new insights into the molecular mechanism of rice panicle structure regulation by BPB1 by regulating lignin and cellulose content and several transcriptional metabolic pathways. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01389-x.

Mapping of qChalk1 controlling grain chalkiness in japonica rice.

BACKGROUND: Rice grain chalkiness is an undesirable characteristic that affects grain quality. The aim of this study was to map QTLs controlling grain chalkiness in japonica rice. METHODS AND RESULTS: In this study, two japonica rice cultivars with similar grain shapes but different grain chalkiness rates were crossed and the F2 and BC1F2 populations were subjected to QTL-seq analysis to map the QTLs controlling the grain chalkiness rate. QTL-seq analysis revealed SNP index differences on chromosome 1 in both of the segregating populations. Using polymorphic markers between the two parents, QTL mapping was conducted on 213 individual plants in the BC1F2 population. QTL mapping confined a QTL controlling grain chalkiness, qChalk1, to a 1.1 Mb genomic region on chromosome 1. qChalk1 explained 19.7% of the phenotypic variation. CONCLUSION: A QTL controlling grain chalkiness qChalk1 was detected in both F2 and BC1F2 segregating populations by QTL-Seq and QTL mapping methods. This result would be helpful for further cloning of the genes controlling grain chalkiness in japonica rice.

Purple Rehmannia: Investigation of the activation of R2R3-MYB transcription factors involved in anthocyanin biosynthesis.

Engineering anthocyanin biosynthesis in herbs could provide health-promoting foods for improving human health. Rehmannia glutinosa is a popular medicinal herb in Asia, and was a health food for the emperors of the Han Dynasty (59 B.C.). In this study, we revealed the differences in anthocyanin composition and content between three Rehmannia species. On the 250, 235 and 206 identified MYBs in the respective species, six could regulate anthocyanin biosynthesis by activating the ANTHOCYANIDIN SYNTHASE (ANS) gene expression. Permanent overexpression of the Rehmannia MYB genes in tobacco strongly promoted anthocyanin content and expression levels of NtANS and other genes. A red appearance of leaves and tuberous/roots was observed, and the total anthocyanin content and the cyanidin-3-O-glucoside content were significantly higher in the lines overexpressing RgMYB41, RgMYB42, and RgMYB43 from R. glutinosa, as well as RcMYB1 and RcMYB3 in R. chingii and RhMYB1 from R. henryi plants. Knocking out of RcMYB3 by CRISPR/Cas9 gene editing resulted in the discoloration of the R. chingii corolla lobes, and decreased the content of anthocyanin. R. glutinosa overexpressing RcMYB3 displayed a distinct purple color in the whole plants, and the antioxidant activity of the transgenic plants was significantly enhanced compared to WT. These results indicate that Rehmannia MYBs can be used to engineer anthocyanin biosynthesis in herbs to improve their additional value, such as increased antioxidant contents.

Proteomic Profiling Reveals the Molecular Control of Oocyte Maturation.

Meiotic maturation is an intricate and precisely regulated process orchestrated by various pathways and numerous proteins. However, little is known about the proteome landscape during oocytes maturation. Here, we obtained the temporal proteomic profiles of mouse oocytes during in vivo maturation. We successfully quantified 4694 proteins from 4500 oocytes in three key stages (germinal vesicle, germinal vesicle breakdown, and metaphase II). In particular, we discovered the novel proteomic features during oocyte maturation, such as the active Skp1-Cullin-Fbox pathway and an increase in mRNA decay-related proteins. Using functional approaches, we further identified the key factors controlling the histone acetylation state in oocytes and the vital proteins modulating meiotic cell cycle. Taken together, our data serve as a broad resource on the dynamics occurring in oocyte proteome and provide important knowledge to better understand the molecular mechanisms during germ cell development.

ZFP57 regulates DNA methylation of imprinted genes to facilitate embryonic development of somatic cell nuclear transfer embryos in Holstein cows.

Aberrant epigenetic nuclear reprogramming, especially imprinting pattern disorders, is one of the major causes of failure of clone development from somatic cell nuclear transfer (SCNT). Previous studies showed that ZFP57 is a key protein required for imprint maintenance after fertilization. In this study, we found that imprinting control regions in several imprinted genes were significantly hypomethylated in cloned embryos compared with in vitro fertilization embryos, indicating a loss of imprinted gene methylation. The ZFP57 expression was capable of maintaining the correct degree of methylation at several imprinting control regions and correcting abnormal hypomethylation. Moreover, we successfully obtained bovine fetal fibroblasts overexpressing ZFP57, which were used as donors for SCNT. Our results demonstrated that overexpression of ZFP57 increased total and trophectoderm cell numbers and the ratio of inner cell mass to total cells, reduced the apoptosis rate and significantly enhanced the development of SCNT blastocysts in vitro, ultimately achieving a degree of methylation similar to that in in vitro fertilization embryos. We concluded that overexpression of ZFP57 in donor cells provided an effective method for enhancing nuclear reprogramming and developmental potential in SCNT embryos. The ZFP57 protein played a key role in maintaining the methylation of imprinted genes during early embryonic development, which may be effective for enhanced SCNT in cattle.

Transcriptome and Metabolome Analyses Reveals the Pathway and Metabolites of Grain Quality Under Phytochrome B in Rice (Oryza sativa L.).

BACKGROUND: Grain size and chalkiness is a critical agronomic trait affecting rice yield and quality. The application of transcriptomics to rice has widened the understanding of complex molecular responsive mechanisms, differential gene expression, and regulatory pathways under varying conditions. Similarly, metabolomics has also contributed drastically for rice trait improvements. As master regulators of plant growth and development, phys influence seed germination, vegetative growth, photoperiodic flowering, shade avoidance responses. OsPHYB can regulate a variety of plant growth and development processes, but little is known about the roles of rice gene OsPHYB in modulating grain development. RESULTS: In this study, rice phytochrome B (OsPHYB) was edited using CRISPR/Cas9 technology. We found that OsPHYB knockout increased rice grain size and chalkiness, and increased the contents of amylose, free fatty acids and soluble sugar, while the gel consistency and contents of proteins were reduced in mutant grains. Furthermore, OsPHYB is involved in the regulation of grain size and chalk formation by controlling cell division and complex starch grain morphology. Transcriptomic analysis revealed that loss of OsPHYB function affects multiple metabolic pathways, especially enhancement of glycolysis, fatty acid, oxidative phosphorylation, and antioxidant pathways, as well as differential expression of starch and phytohormone pathways. An analysis of grain metabolites showed an increase in the free fatty acids and lysophosphatidylcholine, whereas the amounts of sugars, alcohols, amino acids and derivatives, organic acids, phenolic acids, alkaloids, nucleotides and derivatives, and flavonoids decreased, which were significantly associated with grain size and chalk formation. CONCLUSIONS: Our study reveals that, OsPHYB plays an important regulatory role in the growth and development of rice grains, especially grain size and chalkiness. Furthermore, OsPHYB regulates grain size and chalkiness formation by affecting gene metabolism interaction network. Thus, this study not only revealed that OsPHYB plays a vital role in regulating grain size and chalkiness of rice but reveal new functions and highlighted the importance and value of OsPHYB in rice grain development and provide a new strategy for yield and quality improvement in rice breeding.

Modification of alternative splicing in bovine somatic cell nuclear transfer embryos using engineered CRISPR-Cas13d.

Animal cloning can be achieved by somatic cell nuclear transfer (SCNT), but the resulting live birth rate is relatively low. We previously improved the efficiency of bovine SCNT by exogenous melatonin treatment or by overexpression of lysine-specific demethylase 4D (KDM4D) and 4E (KDM4E). In this study, we revealed abundant alternative splicing (AS) transitions during fertilization and embryonic genome activation, and demonstrated abnormal AS in bovine SCNT embryos compared with in vitro fertilized embryos. We used the CRISPR-Cas13d RNA-targeting system to target cis-elements of ABI2 and ZNF106 pre-mRNA to modify AS, thus reducing the ratio of abnormal-isoform SCNT embryos by nearly 50% and achieving a high survival rate (11%-19%). These results indicate that this system may provide an efficient method for bovine cloning, while also paving the way for further improvements in the efficiency of SCNT.

Prism of Employee Performance Through the Means of Internal Support: A Study of Perceived Organizational Support.

The purpose of this paper is to develop a theoretical framework for testing the perceived organizational support (POS)-employee performance relationship with a mediating effect of intrinsic motivation. To combine the concepts from perceived organizational support (POS) and employee performance, secondary data have been collected from different research papers to provide a literature-based analysis. The present study found positive relationships between POS, intrinsic motivation, and employees' performance. The study further realized the intrinsic motivation as a potential mediator between POS and performance relationship. From this framework, numerous areas of research can be pursued to be used to research and practice human resource management. The theoretical framework which is developed in this paper is based upon literature that can be proved empirically. To improve theory, research, and practice in the field of human resource management, this paper conceptualizes the concept of POS through the means of internal support.

OsCKX2 regulates phosphate deficiency tolerance by modulating cytokinin in rice.

Cytokinin oxidase/dehydrogenases (CKXs) are key enzymes that degrade cytokinins (CTKs) and play an essential role in plant growth and development. The present study analyzed the phenotypic and physiological characteristics of OsCKX2 overexpressing (OE) and knockout (KO) rice plants after exposure to phosphate (Pi) deficiency and the transcriptome and metabolome to investigate the function of OsCKX2 in response to Pi deficiency. OsCKX2 KO plants demonstrated higher endogenous CTK levels than wild-type (WT) under Pi deficiency. Further analysis indicated more robust tolerance of OsCKX2 KO plants to Pi deficiency, which exhibited higher phosphorus concentration, larger shoot biomass, and lesser leaf yellowing under Pi deficiency; whereas the opposite was observed for OsCKX2 OE plants. Transcriptome and metabolome analyses revealed that overexpression of OsCKX2 downregulated the transcriptional levels of genes related to Pi transporters, membrane lipid metabolism, and glycolysis, and reduced the consumption of metabolites in membrane lipid metabolism and glycolysis. On the contrary, knockout of OsCKX2 upregulated the expression of Pi transporters, and increased the consumption of metabolites in membrane lipid metabolism and glycolysis. These results indicated that OsCKX2 impacted Pi uptake, recycling, and plant growth via Pi transporters, phospholipid hydrolysis, and glycolysis under Pi deficiency. Overall, OsCKX2 negatively regulated Pi deficiency tolerance by modulating CTKs in rice.

Regulation of OsPIL15 on rice quality.

The phytochrome-interacting factor-like gene OsPIL15 negatively regulates grain size and 1000-grain weight, but its regulatory effect on rice quality traits is unknown. Here, knock-down, knock-out, and over-expression of OsPIL15 transgenic rice lines were used to investigate the effects of OsPIL15 on rice yield and quality traits. The results showed that knock-down or knock-out of OsPIL15 increased grain length and width, chalkiness, amylose content, glutenin and globulin content, and total protein content but reduced amylopectin content, total starch content, prolamin and albumin content, and gel consistency. Over-expression of OsPIL15 showed the opposite results, except for the reduction of prolamin content. Although OsPIL15 changed the grain size and weight, it had no effect on grain length/width ratio, brown rice rate, and milled rice rate. KEGG pathway enrichment analysis of differentially expressed genes between transgenic lines and wild type showed that OsPIL15 mainly regulated genes related to ribosome, metabolic pathways, and biosynthesis of secondary metabolites. Gene expression analysis showed that RNAi transgenic lines decreased OsCIN2 and OsSUS1 expression and increased OsGBSSI, OsSSI, OsAPGL2, and OsAPGL3 expression level, while over-expression of OsPIL15 increased OsCIN2, OsSUS1, OsSUS6, and OsSSI and decreased OsSSIIa, OsSSIIc, and OsAPGL2 expression level. These results revealed that OsPIL15 plays an important role in rice grain development. In addition to grain shape, OsPIL15 also regulates chalkiness, starch content, protein content, and gel consistency. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01311-x.

CHAF1b, chromatin assembly factor-1 subunit b, is essential for mouse preimplantation embryos.

Chromatin assembly factor-1, subunit b (CHAF1b), the p60 subunit of the chromatin-assembly factor-1 (CAF-1) complex, is an evolutionarily conserved protein that has been implicated in various biological processes. Although a variety of functions have been attributed to CHAF1b, its function in preimplantation embryos remains obscure. In this study, we showed that CHAF1b knockdown did not affect the blastocyst rate, but resulted in a low blastocyst hatching rate, outgrowth failure in vitro, and embryonic lethality after implantation in vivo. Notably, CHAF1b depletion increased apoptosis and caused down-regulated expression of key regulators of cell fate specification, including Oct4, Cdx2, Sox2, and Nanog. Further analysis revealed that CHAF1b mediated the replacement of H3.3 with H3.1/3.2, which was associated with decreased repressive histone marks (H3K9me2/3 and H3K27me2/3) and increased active histone marks (H3K4me2/3). Moreover, RNA-sequencing analysis revealed that CHAF1b depletion resulted in the differential expression of 1508 genes, including epigenetic modifications genes, multiple lineage-specific genes, and several genes encoding apoptosis proteins. In addition, assay for transposase-accessible chromatin-sequencing analysis demonstrated that silencing CHAF1b altered the chromatin accessibility of lineage-specific genes and epigenetic modifications genes. Taken together, these data imply that CHAF1b plays significant roles in preimplantation embryos, probably by regulating epigenetic modifications and lineage specification.

Repellent, larvicidal and adulticidal activities of essential oil from Dai medicinal plant Zingiber cassumunar against Aedes albopictus.

Zingiber cassumunar is an important plant used in traditional medicine and as a natural mosquito repellent. However, the compounds responsible for the repellent activity of the plant are still unknown. The aim of the study is to identify the components of Z. cassumunar essential oil that show repellent activity against Aedes albopictus. We also evaluated the larvicidal and adulticidal activities of Z. cassumunar essential oil against Ae. albopictus. In-cage mosquito repellent experiments showed that Z. cassumunar essential oil possessed moderate repellent activity with a minimum effective dose (MED) of 0.16 ± 0.01 mg/cm2, compared to reference standard N,N-diethyl-3-methylbenzamide (DEET, 0.03 ± 0.01 mg/cm2). Bioassay-guided fractionation identified the major active compound of Z. cassumunar essential oil as (-)-terpinen-4-ol (1) (MED: 0.19 ± 0 mg/cm2). We also found that Z. cassumunar essential oil showed moderate larvicidal activity against first instar larvae of Ae. albopictus with a LC50 (50% lethal concentration) of 44.9 µg/L after 24 h. Fumigation bioassays showed that Z. cassumunar essential oil exhibits moderate adulticidal activity against Ae. albopictus with a LC50 of 5.44%, while (-)-terpinen-4-ol showed significant adulticidal activity with a LC50 of 2.10% after 24 h. This study verifies that the Z. cassumunar essential oil has mosquito repellent activity, and that (-)-terpinen-4-ol is mainly responsible for this activity. Furthermore, this study provides scientific support for the folk usage of Z. cassumunar essential oil as mosquito repellent and indicates that Z. cassumunar essential oil and (-)-terpinen-4-ol can be used as plant-derived repellents and insecticides for mosquito control.

Efficient CRISPR/Cas9-mediated genome editing in Rehmannia glutinosa.

KEY MESSAGE: Here, we cloned a phytoene desaturase (PDS) gene from Rehmannia glutinosa, and realized RgPDS1 knock out in R. glutinosa resulted in the generation of albino plants. Rehmannia glutinosa is a highly important traditional Chinese medicine (TCM) with specific pharmacology and economic value. R. glutinosa is a tetraploid plant, to date, no report has been published on gene editing of R. glutinosa. In this study, we combined the transcriptome database of R. glutinosa and the reported phytoene desaturase (PDS) gene sequences to obtain the PDS gene of R. glutinosa. Then, the PDS gene was used as a marker gene to verify the applicability and gene editing efficiency of the CRISPR/Cas9 system in R. glutinosa. The constructed CRISPR/Cas9 system was mediated by Agrobacterium to genetically transform into R. glutinosa, and successfully regenerated fully albino and chimeric albino plants. The next-generation sequencing (NGS) confirmed that the albino phenotype was indeed caused by RgPDS gene target site editing, and it was found that base deletion was more common than insertion or replacement. Our results revealed that zCas9 has a high editing efficiency on the R. glutinosa genome. This research lays a foundation for further use of gene editing technology to study the molecular functions of genes, create excellent germplasm, accelerate domestication, and improve the yield and quality of R. glutinosa.