An integrated rice panicle phenotyping method based on X-ray and RGB scanning and deep learning

Rice panicle phenotyping is required in rice breeding for high yield and grain quality.To fully evaluate spikelet and kernel traits without threshing and hulling,using X-ray and RGB scanning,we developed an integrated rice panicle phenotyping system and a corresponding image analysis pipeline.We compared five methods of counting spikelets and found that Faster R-CNN achieved high accuracy(R~2 of 0.99)and speed.Faster R-CNN was also applied to indica and japonica classification and achieved 91%accuracy.The proposed integrated panicle phenotyping method offers benefit for rice functional genetics and breeding.

Rice Functional Genomics Research： Past Decade and Future

Rice （Oryza sativa） is a major staple food crop for more than 3.5 billion people worldwide. Under- standing the regulatory mechanisms of complex agronomic traits in rice is critical for global food security. Rice is also a model plant for genomics research of monocotyledonso Thanks to the rapid development of functional genomic technologies, over 2000 genes controlling important agronomic traits have been cloned, and their molecular biological mechanisms have also been partially char- acterized. Here, we briefly review the advances in rice functional genomics research during the past 10 years, including a summary of functional genomics platforms, genes and molecular regulatory networks that regulate important agronomic traits, and newly developed tools for gene identification. These achievements made in functional genomics research will greatly facilitate the development of green super rice. We also discuss future challenges and prospects of rice functional genomics research.

Effort and Contribution of T-DNA Insertion Mutant Library for Rice Functional Genomics Research in China:Review and Perspective

With the completion of the rice （Oryza sativa L.） genome-sequencing project, the rice research community proposed to characterize the func- tion of every predicted gene in rice by 2020. One of the most effective and high-throughput strategies for studying gene function is to employ genetic mutations induced by insertion elements such as T-DNA or transposons. Since 1999, with support from the Ministry of Science and Technology of China for Rice Functional Genomics Programs, large-scale T-DNA insertion mutant populations have been generated in Huazhong Agricultural University, the Chinese Academy of Sciences and the Chinese Academy of Agricultural Sciences. Currently, a total of 372,346 mutant lines have been generated, and 58,226 T-DNA or Tos17 flanking sequence tags have been isolated. Using these mutant resources, more than 40 genes with potential applications in rice breeding have already been identified. These include genes involved in biotic or abiotic stress responses, nutrient metabolism, pollen development, and plant architecture. The functional analysis of these genes will not only deepen our understanding of the fundamental biological questions in rice, but will also offer valuable gene resources for developing Green Super Rice that is high-yielding with few inputs even under the poor growth conditions of many regions of Africa and Asia.

Phosphorylation of OsFD1 by OsCIPK3 promotes the formation of RFT1-containing florigen activation complex for long-day flowering in rice

Heading date is a critical trait that determines the regional adaptability and grain productivity of many crops.Although rice is a facultative short-day plant,its domestication led to the Ghd7-Ehd1-Hd3a/RFT1 pathway for adaptation to long-day conditions(LDs).The formation of the"florigen activation complex"(FAC)containing florigen Hd3a has been characterized.However,the molecular composition of the FAC that contains RFT1 for long-day flowering is unclear.We show here that RFT1 forms a ternary FAC with 14-3-3 proteins and OsFD1 to promote flowering under LDs.We identified a calcineurin B-like-interacting protein kinase,OsCIPK3,which directly interacts with and phosphorylates OsFD1,thereby facilitating the localization of the FAC to the nucleus.Mutation in OsCIPK3 results in a late heading date under LDs but a normal heading date under short-day conditions.Collectively,our results suggest that OsCIPK3 phosphorylates OsFD1 to promote RFT1-containing FAC formation and consequently induce flowering in rice under LDs.

miR395-regulated sulfate metabolism exploits pathogen sensitivity to sulfate to boost immunity in rice

MicroRNAs (miRNAs) play important roles in plant physiological activities. However, their roles and molecular mechanisms in boosting plant immunity, especially through the modulation of macronutrient metabolism in response to pathogens, are largely unknown. Here, we report that an evolutionarily conserved miRNA, miR395, promotes resistance to Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc), two destructive bacterial pathogens, by regulating sulfate accumulation and distribution in rice. Specifically, miR395 targets and suppresses the expression of the ATP sulfurylase gene OsAPS1, which functions in sulfate assimilation, and two sulfate transporter genes, OsSULTR2;1 and OsSULTR2;2, which function in sulfate translocation, to promote sulfate accumulation, resulting in broad-spectrum resistance to bacterial pathogens in miR395-overexpressing plants. Genetic analysis revealed that miR395-triggered resistance is involved in both pathogen-associated molecular pattern-triggered immunity and R gene-mediated resistance. Moreover, we found that accumulated sulfate but not S-metabolites inhibits proliferation of pathogenic bacteria, revealing a sulfate-mediated antibacterial defense mechanism that differs from sulfur-induced resistance. Furthermore, compared with other bacteria, Xoo and Xoc, which lack the sulfate transporter CysZ, are sensitive to high levels of extracellular sulfate. Accordingly, miR395-regulated sulfate accumulation impaired the virulence of Xoo and Xoc by decreasing extracellular polysaccharide production and biofilm formation. Taken together, these results suggest that rice miR395 modulates sulfate metabolism to exploit pathogen sensitivity to sulfate and thereby promotes broad-spectrum resistance.

Genome-wide binding analysis of transcription factor Rice Indeterminate 1 reveals a complex network controlling rice floral transition

RICE INDETERMINATE 1(RID1)plays a critical role in controlling floral transition in rice(Oryza sativa).However,the molecular basis for this effect,particularly the target genes and regulatory specificity,remains largely unclear.Here,we performed chromatin immunoprecipitation followed by sequencing(ChIP-seq)in young leaves at the pre-floral-transition stage to identify the target genes of RID1,identifying 2,680 genes associated with RID1 binding sites genome-wide.RID1 binding peaks were highly enriched for TTTGTC,the direct binding motif of the INDETERMINATE DOMAIN protein family that includes RID1.Interestingly,CACGTG and GTGGGCCC,two previously uncharacterized indirect binding motifs,were enriched through the interactions of RID1 with the novel floweringpromoting proteins OsPIL12 and OsTCP11,respectively.Moreover,the ChIP-seq data demonstrated that RID1 bound to numerous rice heading-date genes,such as HEADING DATE1(HD1)and FLAVIN-BINDING,KELCH REPEAT,F-BOX 1(OsFKF1).Notably,transcriptome sequencing(RNA-seq)analysis revealed roles of RID1 in diverse developmental pathways.Genetic analysis combined with genome-wide ChIP-seq and RNA-seq results showed that RID1 directly binds to the promoter of OsERF#136(a repressor of rice flowering)and negatively regulates its expression.Overall,our findings provide new insights into the molecular and genetic mechanisms underlying rice floral transition and characterize OsERF#136 as a previously unrecognized direct target of RID1.

OsMLH1 interacts with OsMLH3 to regulate synapsis and interference-sensitive crossover formation during meiosis in rice

Meiotic recombination is essential for reciprocal exchange of genetic information between homologous chromosomes and their subsequent proper segregation in sexually reproducing organisms. MLH1 and MLH3 belong to meiosis-specific members of the Mut L-homolog family, which are required for normal level of crossovers(COs) in some eukaryotes. However, their functions in plants need to be further elucidated.Here, we report the identification of Os MLH1 and reveal its functions during meiosis in rice. Using CRISPRCas9 approach, two independent mutants, Osmlh1-1 and Osmlh1-2, are generated and exhibited significantly reduced male fertility. In Osmlh1-1, the clearance of PAIR2 is delayed and partial ZEP1 proteins are not loaded into the chromosomes, which might be due to the deficient in resolution of interlocks at late zygotene. Thus, Os MLH1 is required for the assembly of synapsis complex. In Osmlh1-1, CO number is dropped by ~53% and the distribution of residual COs is consistent with predicted Poisson distribution,indicating that Os MLH1 is essential for the formation of interference-sensitive COs(class I COs). Os MLH1 interacts with Os MLH3 through their C-terminal domains. Mutation in Os MLH3 also affects the pollen fertility. Thus, our experiments reveal that the conserved heterodimer Mut Lg(Os MLH1-Os MLH3) is essential for the formation of class I COs in rice.