Plant microphenotype: from innovative imaging to computational analysis.

The microphenotype plays a key role in bridging the gap between the genotype and the complex macro phenotype. In this article, we review the advances in data acquisition and the intelligent analysis of plant microphenotyping and present applications of microphenotyping in plant science over the past two decades. We then point out several challenges in this field and suggest that cross-scale image acquisition strategies, powerful artificial intelligence algorithms, advanced genetic analysis, and computational phenotyping need to be established and performed to better understand interactions among genotype, environment, and management. Microphenotyping has entered the era of Microphenotyping 3.0 and will largely advance functional genomics and plant science.

Phenotype identification and genome-wide association study of ear-internode vascular bundles in maize (Zea mays).

The vascular bundle in the ear-internode of maize is a key conduit for transporting photosynthetic materials between "source" and "sink", making it critically important to examine its micro-phenotypes and genetic architecture to identify advantageous characteristics and cultivate high-yielding and high-quality varieties. Unfortunately, the limited observation methods and scope of study precludes any comprehensive and systematic investigations into the microscopic phenotypes and genetic mechanisms of vascular bundle in maize ear-internode. In this study, 47 phenotypic traits were extracted in 495 maize inbred lines using micro computed tomography (Micro-CT) scanning technology and a deep learning-based phenotype acquisition method for stem vascular bundle, which included stem slice-related, epidermis zone-related, periphery zone-related, inner zone-related and vascular bundles-related traits. Phenotypic analysis indicated that there was extensive phenotypic variation of vascular bundle traits in ear-internode, especially that in the inner zone. Of these, 30 phenotypic traits with heritability greater than 0.70 were conducted for GWAS, and a total of 4,225 significant SNPs and 416 candidate genes with detailed functional annotations were identified. Furthermore, 20 genes were highly expressed in stem-related tissues, especially in maize internodes. Functional analysis of candidate genes indicated that the pathways obtained for candidate genes of different trait groups were distinct, mainly involved in vitamin synthesis and metabolism, transport of substances, carbohydrate derivative catabolic process, protein transport and localization, and anatomical structure development. The results of this study will help to further understand the phenotypic traits of stem vascular bundles and provide a reference for revealing the genetic mechanism of maize ear-internode vascular bundles.

Dissecting the phenotypic components and genetic architecture of maize stem vascular bundles using high-throughput phenotypic analysis.

High-throughput phenotyping is increasingly becoming an important tool for rapid advancement of genetic gain in breeding programmes. Manual phenotyping of vascular bundles is tedious and time-consuming, which lags behind the rapid development of functional genomics in maize. More robust and automated techniques of phenotyping vascular bundles traits at high-throughput are urgently needed for large crop populations. In this study, we developed a standard process for stem micro-CT data acquisition and an automatic CT image process pipeline to obtain vascular bundle traits of stems including geometry-related, morphology-related and distribution-related traits. Next, we analysed the phenotypic variation of stem vascular bundles between natural population subgroup (480 inbred lines) based on 48 comprehensively phenotypic information. Also, the first database for stem micro-phenotypes, MaizeSPD, was established, storing 554 pieces of basic information of maize inbred lines, 523 pieces of experimental information, 1008 pieces of CT scanning images and processed images, and 24 192 pieces of phenotypic data. Combined with genome-wide association studies (GWASs), a total of 1562 significant single nucleotide polymorphism (SNPs) were identified for 30 stem micro-phenotypic traits, and 84 unique genes of 20 traits such as VBNum, VBAvArea and PZVBDensity were detected. Candidate genes identified by GWAS mainly encode enzymes involved in cell wall metabolism, transcription factors, protein kinase and protein related to plant signal transduction and stress response. The results presented here will advance our knowledge about phenotypic trait components of stem vascular bundles and provide useful information for understanding the genetic controls of vascular bundle formation and development.

Research on Recognition of Motor Imagination Based on Connectivity Features of Brain Functional Network.

Feature extraction is essential for classifying different motor imagery (MI) tasks in a brain-computer interface. To improve classification accuracy, we propose a novel feature extraction method in which the connectivity increment rate (CIR) of the brain function network (BFN) is extracted. First, the BFN is constructed on the basis of the threshold matrix of the Pearson correlation coefficient of the mu rhythm among the channels. In addition, a weighted BFN is constructed and expressed by the sum of the existing edge weights to characterize the cerebral cortex activation degree in different movement patterns. Then, on the basis of the topological structures of seven mental tasks, three regional networks centered on the C3, C4, and Cz channels are constructed, which are consistent with correspondence between limb movement patterns and cerebral cortex in neurophysiology. Furthermore, the CIR of each regional functional network is calculated to form three-dimensional vectors. Finally, we use the support vector machine to learn a classifier for multiclass MI tasks. Experimental results show a significant improvement and demonstrate the success of the extracted feature CIR in dealing with MI classification. Specifically, the average classification performance reaches 88.67% which is higher than other competing methods, indicating that the extracted CIR is effective for MI classification.

High-throughput micro-phenotyping measurements applied to assess stalk lodging in maize (Zea mays L.).

BACKGROUND: The biomechanical properties of maize stalks largely determine their lodging resistance, which affects crop yield per unit area. However, the quantitative and qualitative relationship between micro-phenotypes and the biomechanics of maize stalks is still under examined. In particular, the roles of the number, geometry, and distribution of vascular bundles of stalks in maize lodging resistance remain unclear. Research on these biomechanical properties will benefit from high-resolution micro-phenotypic image acquisition capabilities, which have been improved by modern X-ray imaging devices such as micro-CT and the development of micro-phenotyping analysis software. Hence, high-throughput image analysis and accurate quantification of anatomical phenotypes of stalks are necessary. RESULTS: We have updated VesselParser version 1.0 to version 2.0 and have improved its performance, accuracy, and computation strategies. Anatomical characteristics of the second and third stalk internodes of the cultivars 'Jingke968' and 'Jingdan38' were analyzed using VesselParser 2.0. The relationships between lodging resistance and anatomical phenotypes of stalks between the two different maize varieties were investigated. The total area of vascular bundles in the peripheral layer, auxiliary axis diameter, and total area of vascular bundles were revealed to have the highest correlation with mechanical properties, and anatomical phenotypes of maize stalk were better predictors of mechanical properties than macro features observed optically from direct measurement, such as diameter and perimeter. CONCLUSIONS: This study demonstrates the utility of VesselParser 2.0 in assessing stalk mechanical properties. The combination of anatomical phenotypes and mechanical behavior research provides unique insights into the problem of stalk lodging, showing that micro phenotypes of vascular bundles are good predictors of maize stalk mechanical properties that may be important indices for the evaluation and identification of the biomechanical properties to improve lodging resistance of future maize varieties.

Three-Dimensional Modeling of Maize Canopies Based on Computational Intelligence.

The 3-dimensional (3D) modeling of crop canopies is fundamental for studying functional-structural plant models. Existing studies often fail to capture the structural characteristics of crop canopies, such as organ overlapping and resource competition. To address this issue, we propose a 3D maize modeling method based on computational intelligence. An initial 3D maize canopy is created using the t-distribution method to reflect characteristics of the plant architecture. The subsequent model considers the 3D phytomers of maize as intelligent agents. The aim is to maximize the ratio of sunlit leaf area, and by iteratively modifying the azimuth angle of the 3D phytomers, a 3D maize canopy model that maximizes light resource interception can be constructed. Additionally, the method incorporates a reflective approach to optimize the canopy and utilizes a mesh deformation technique for detecting and responding to leaf collisions within the canopy. Six canopy models of 2 varieties plus 3 planting densities was constructed for validation. The average R2 of the difference in azimuth angle between adjacent leaves is 0.71, with a canopy coverage error range of 7% to 17%. Another 3D maize canopy model constructed using 12 distinct density gradients demonstrates the proportion of leaves perpendicular to the row direction increases along with the density. The proportion of these leaves steadily increased after 9 × 104 plants ha-1. This study presents a 3D modeling method for the maize canopy. It is a beneficial exploration of swarm intelligence on crops and generates a new way for exploring efficient resources utilization of crop canopies.

Using high-throughput phenotype platform MVS-Pheno to reconstruct the 3D morphological structure of wheat.

It is of great significance to study the plant morphological structure for improving crop yield and achieving efficient use of resources. Three dimensional (3D) information can more accurately describe the morphological and structural characteristics of crop plants. Automatic acquisition of 3D information is one of the key steps in plant morphological structure research. Taking wheat as the research object, we propose a point cloud data-driven 3D reconstruction method that achieves 3D structure reconstruction and plant morphology parameterization at the phytomer scale. Specifically, we use the MVS-Pheno platform to reconstruct the point cloud of wheat plants and segment organs through the deep learning algorithm. On this basis, we automatically reconstructed the 3D structure of leaves and tillers and extracted the morphological parameters of wheat. The results show that the semantic segmentation accuracy of organs is 95.2%, and the instance segmentation accuracy AP50 is 0.665. The R2 values for extracted leaf length, leaf width, leaf attachment height, stem leaf angle, tiller length, and spike length were 0.97, 0.80, 1.00, 0.95, 0.99, and 0.95, respectively. This method can significantly improve the accuracy and efficiency of 3D morphological analysis of wheat plants, providing strong technical support for research in fields such as agricultural production optimization and genetic breeding.

Design and Development of a Low-Cost UGV 3D Phenotyping Platform with Integrated LiDAR and Electric Slide Rail.

Unmanned ground vehicles (UGV) have attracted much attention in crop phenotype monitoring due to their lightweight and flexibility. This paper describes a new UGV equipped with an electric slide rail and point cloud high-throughput acquisition and phenotype extraction system. The designed UGV is equipped with an autopilot system, a small electric slide rail, and Light Detection and Ranging (LiDAR) to achieve high-throughput, high-precision automatic crop point cloud acquisition and map building. The phenotype analysis system realized single plant segmentation and pipeline extraction of plant height and maximum crown width of the crop point cloud using the Random sampling consistency (RANSAC), Euclidean clustering, and k-means clustering algorithm. This phenotyping system was used to collect point cloud data and extract plant height and maximum crown width for 54 greenhouse-potted lettuce plants. The results showed that the correlation coefficient (R2) between the collected data and manual measurements were 0.97996 and 0.90975, respectively, while the root mean square error (RMSE) was 1.51 cm and 4.99 cm, respectively. At less than a tenth of the cost of the PlantEye F500, UGV achieves phenotypic data acquisition with less error and detects morphological trait differences between lettuce types. Thus, it could be suitable for actual 3D phenotypic measurements of greenhouse crops.

Disentangling the Heterosis in Biomass Production and Radiation Use Efficiency in Maize: A Phytomer-Based 3D Modelling Approach.

Maize (Zea mays L.) benefits from heterosis in-yield formation and photosynthetic efficiency through optimizing canopy structure and improving leaf photosynthesis. However, the role of canopy structure and photosynthetic capacity in determining heterosis in biomass production and radiation use efficiency has not been separately clarified. We developed a quantitative framework based on a phytomer-based three-dimensional canopy photosynthesis model and simulated light capture and canopy photosynthetic production in scenarios with and without heterosis in either canopy structure or leaf photosynthetic capacity. The accumulated above-ground biomass of Jingnongke728 was 39% and 31% higher than its male parent, Jing2416, and female parent, JingMC01, while accumulated photosynthetically active radiation was 23% and 14% higher, correspondingly, leading to an increase of 13% and 17% in radiation use efficiency. The increasing post-silking radiation use efficiency was mainly attributed to leaf photosynthetic improvement, while the dominant contributing factor differs for male and female parents for heterosis in post-silking yield formation. This quantitative framework illustrates the potential to identify the key traits related to yield and radiation use efficiency and helps breeders to make selections for higher yield and photosynthetic efficiency.

Geometric Wheat Modeling and Quantitative Plant Architecture Analysis Using Three-Dimensional Phytomers.

The characterization, analysis, and evaluation of morphology and structure are crucial in wheat research. Quantitative and fine characterization of wheat morphology and structure from a three-dimensional (3D) perspective has great theoretical significance and application value in plant architecture identification, high light efficiency breeding, and cultivation. This study proposes a geometric modeling method of wheat plants based on the 3D phytomer concept. Specifically, 3D plant architecture parameters at the organ, phytomer, single stem, and individual plant scales were extracted based on the geometric models. Furthermore, plant architecture vector (PA) was proposed to comprehensively evaluate wheat plant architecture, including convergence index (C), leaf structure index (L), phytomer structure index (PHY), and stem structure index (S). The proposed method could quickly and efficiently achieve 3D wheat plant modeling by assembling 3D phytomers. In addition, the extracted PA quantifies the plant architecture differences in multi-scales among different cultivars, thus, realizing a shift from the traditional qualitative to quantitative analysis of plant architecture. Overall, this study promotes the application of the 3D phytomer concept to multi-tiller crops, thereby providing a theoretical and technical basis for 3D plant modeling and plant architecture quantification in wheat.

Using High-Throughput Phenotyping Analysis to Decipher the Phenotypic Components and Genetic Architecture of Maize Seedling Salt Tolerance.

Soil salinization is a worldwide problem that limits agricultural production. It is important to understand the salt stress tolerance ability of maize seedlings and explore the underlying related genetic resources. In this study, we used a high-throughput phenotyping platform with a 3D laser sensor (Planteye F500) to identify the digital biomass, plant height and normalized vegetation index under normal and saline conditions at multiple time points. The result revealed that a three-leaf period (T3) was identified as the key period for the phenotypic variation in maize seedlings under salt stress. Moreover, we mapped the salt-stress-related SNPs and identified candidate genes in the natural population via a genome-wide association study. A total of 44 candidate genes were annotated, including 26 candidate genes under normal conditions and 18 candidate genes under salt-stressed conditions. This study demonstrates the feasibility of using a high-throughput phenotyping platform to accurately, continuously quantify morphological traits of maize seedlings in different growing environments. And the phenotype and genetic information of this study provided a theoretical basis for the breeding of salt-resistant maize varieties and the study of salt-resistant genes.

Investigating the genetic basis of maize ear characteristics: a comprehensive genome-wide study utilizing high-throughput phenotypic measurement method and system.

The morphology of maize ears plays a critical role in the breeding of new varieties and increasing yield. However, the study of traditional ear-related traits alone can no longer meet the requirements of breeding. In this study, 20 ear-related traits, including size, shape, number, and color, were obtained in 407 maize inbred lines at two sites using a high-throughput phenotypic measurement method and system. Significant correlations were found among these traits, particularly the novel trait ear shape (ES), which was correlated with traditional traits: kernel number per row and kernel number per ear. Pairwise comparison tests revealed that the inbred lines of tropical-subtropical were significantly different from other subpopulations in row numbers per ear, kernel numbers per ear, and ear color. A genome-wide association study identified 275, 434, and 362 Single nucleotide polymorphisms (SNPs) for Beijing, Sanya, and best linear unbiased prediction scenarios, respectively, explaining 3.78% to 24.17% of the phenotypic variance. Furthermore, 58 candidate genes with detailed functional descriptions common to more than two scenarios were discovered, with 40 genes being associated with color traits on chromosome 1. After analysis of haplotypes, gene expression, and annotated information, several candidate genes with high reliability were identified, including Zm00001d051328 for ear perimeter and width, zma-MIR159f for ear shape, Zm00001d053080 for kernel width and row number per ear, and Zm00001d048373 for the blue color channel of maize kernels in the red-green-blue color model. This study emphasizes the importance of researching novel phenotypic traits in maize by utilizing high-throughput phenotypic measurements. The identified genetic loci enrich the existing genetic studies related to maize ears.

Research on the evolutionary history of the morphological structure of cotton seeds: a new perspective based on high-resolution micro-CT technology.

Cotton (Gossypium hirsutum L.) seed morphological structure has a significant impact on the germination, growth and quality formation. However, the wide variation of cotton seed morphology makes it difficult to achieve quantitative analysis using traditional phenotype acquisition methods. In recent years, the application of micro-CT technology has made it possible to analyze the three-dimensional morphological structure of seeds, and has shown technical advantages in accurate identification of seed phenotypes. In this study, we reconstructed the seed morphological structure based on micro-CT technology, deep neural network Unet-3D model, and threshold segmentation methods, extracted 11 basics phenotypes traits, and constructed three new phenotype traits of seed coat specific surface area, seed coat thickness ratio and seed density ratio, using 102 cotton germplasm resources with clear year characteristics. Our results show that there is a significant positive correlation (P< 0.001) between the cotton seed size and that of the seed kernel and seed coat volume, with correlation coefficients ranging from 0.51 to 0.92, while the cavity volume has a lower correlation with other phenotype indicators (r<0.37, P< 0.001). Comparison of changes in Chinese self-bred varieties showed that seed volume, seed surface area, seed coat volume, cavity volume and seed coat thickness increased by 11.39%, 10.10%, 18.67%, 115.76% and 7.95%, respectively, while seed kernel volume, seed kernel surface area and seed fullness decreased by 7.01%, 0.72% and 16.25%. Combining with the results of cluster analysis, during the hundred-year cultivation history of cotton in China, it showed that the specific surface area of seed structure decreased by 1.27%, the relative thickness of seed coat increased by 8.70%, and the compactness of seed structure increased by 50.17%. Furthermore, the new indicators developed based on micro-CT technology can fully consider the three-dimensional morphological structure and cross-sectional characteristics among the indicators and reflect technical advantages. In this study, we constructed a microscopic phenotype research system for cotton seeds, revealing the morphological changes of cotton seeds with the year in China and providing a theoretical basis for the quantitative analysis and evaluation of seed morphology.

Multi-Source Data Fusion Improves Time-Series Phenotype Accuracy in Maize under a Field High-Throughput Phenotyping Platform.

The field phenotyping platforms that can obtain high-throughput and time-series phenotypes of plant populations at the 3-dimensional level are crucial for plant breeding and management. However, it is difficult to align the point cloud data and extract accurate phenotypic traits of plant populations. In this study, high-throughput, time-series raw data of field maize populations were collected using a field rail-based phenotyping platform with light detection and ranging (LiDAR) and an RGB (red, green, and blue) camera. The orthorectified images and LiDAR point clouds were aligned via the direct linear transformation algorithm. On this basis, time-series point clouds were further registered by the time-series image guidance. The cloth simulation filter algorithm was then used to remove the ground points. Individual plants and plant organs were segmented from maize population by fast displacement and region growth algorithms. The plant heights of 13 maize cultivars obtained using the multi-source fusion data were highly correlated with the manual measurements (R2 = 0.98), and the accuracy was higher than only using one source point cloud data (R2 = 0.93). It demonstrates that multi-source data fusion can effectively improve the accuracy of time series phenotype extraction, and rail-based field phenotyping platforms can be a practical tool for plant growth dynamic observation of phenotypes in individual plant and organ scales.

Three-dimensional branch segmentation and phenotype extraction of maize tassel based on deep learning.

BACKGROUND: The morphological structure phenotype of maize tassel plays an important role in plant growth, reproduction, and yield formation. It is an important step in the distinctness, uniformity, and stability (DUS) testing to obtain maize tassel phenotype traits. Plant organ segmentation can be achieved with high-precision and automated acquisition of maize tassel phenotype traits because of the advances in the point cloud deep learning method. However, this method requires a large number of data sets and is not robust to automatic segmentation of highly adherent organ components; thus, it should be combined with point cloud processing technology. RESULTS: An innovative method of incomplete annotation of point cloud data was proposed for easy development of the dataset of maize tassels,and an automatic maize tassel phenotype analysis system: MaizeTasselSeg was developed. The tip feature of point cloud is trained and learned based on PointNet + + network, and the tip point cloud of tassel branch was automatically segmented. Complete branch segmentation was realized based on the shortest path algorithm. The Intersection over Union (IoU), precision, and recall of the segmentation results were 96.29, 96.36, and 93.01, respectively. Six phenotypic traits related to morphological structure (branch count, branch length, branch angle, branch curvature, tassel volume, and dispersion) were automatically extracted from the segmentation point cloud. The squared correlation coefficients (R2) for branch length, branch angle, and branch count were 0.9897, 0.9317, and 0.9587, respectively. The root mean squared error (RMSE) for branch length, branch angle, and branch count were 0.529 cm, 4.516, and 0.875, respectively. CONCLUSION: The proposed method provides an efficient scheme for high-throughput organ segmentation of maize tassels and can be used for the automatic extraction of phenotypic traits of maize tassel. In addition, the incomplete annotation approach provides a new idea for morphology-based plant segmentation.

Prediction of vertical distribution of SPAD values within maize canopy based on unmanned aerial vehicles multispectral imagery.

Real-time monitoring of canopy chlorophyll content is significant in understanding crop growth status and guiding precision agricultural management. Remote sensing methods have demonstrated great potential in this regard. However, the spatiotemporal heterogeneity of chlorophyll content within crop canopies poses challenges to the accuracy and stability of remote sensing estimation models. Hence, this study aimed to develop a novel method for estimating canopy chlorophyll content (represented by SPAD values) in maize (Zea mays L.) canopies. Firstly, we investigated the spatiotemporal distribution patterns of maize canopy SPAD values under varying nitrogen application rates and different growth stages. The results revealed a non-uniform, "bell-shaped" curve distribution of maize canopy SPAD values in the vertical direction. Nitrogen application significantly influenced the distribution structure of SPAD values within the canopy. Secondly, we achieved satisfactory results by fitting the Lorentz peak distribution function to the SPAD values of different leaf positions in maize. The fitting performance, evaluated using R2 and RMSE, ranged from 0.69 to 0.98 and 0.45 to 3.59, respectively, for the year 2021, and from 0.69 to 0.77 and 2.38 to 6.51, respectively, for the year 2022.Finally, based on the correlation between canopy SPAD values and vegetation indices (VIs) at different growth stages, we identified the sensitive leaf positions for the selected CCCI (Canopy Chlorophyll Index) in each growth stage. The 6th (r = 0.662), 4th (r = 0.816), 12th (r = 0.722), and 12th (r = 0.874) leaf positions exhibited the highest correlations. Compared to the estimation model using canopy wide SPAD values, the model based on sensitive leaf positions showed improved accuracy, with increases of 34%, 3%, 20%, and 3% for each growth stage, respectively. In conclusion, the findings of this study contribute to the enhancement of chlorophyll content estimation models in crop canopies and provide valuable insights for the integration of crop growth models with remote sensing methods.

LettuceGDB: The community database for lettuce genetics and omics.

As a globally popular leafy vegetable and a representative plant of the Asteraceae family, lettuce has great economic and academic significance. In the last decade, high-throughput sequencing, phenotyping, and other multi-omics data in lettuce have accumulated on a large scale, thus increasing the demand for an integrative lettuce database. Here, we report the establishment of a comprehensive lettuce database, LettuceGDB (https://www.lettucegdb.com/). As an omics data hub, the current LettuceGDB includes two reference genomes with detailed annotations; re-sequencing data from over 1000 lettuce varieties; a collection of more than 1300 worldwide germplasms and millions of accompanying phenotypic records obtained with manual and cutting-edge phenomics technologies; re-analyses of 256 RNA sequencing datasets; a complete miRNAome; extensive metabolite information for representative varieties and wild relatives; epigenetic data on the genome-wide chromatin accessibility landscape; and various lettuce research papers published in the last decade. Five hierarchically accessible functions (Genome, Genotype, Germplasm, Phenotype, and O-Omics) have been developed with a user-friendly interface to enable convenient data access. Eight built-in tools (Assembly Converter, Search Gene, BLAST, JBrowse, Primer Design, Gene Annotation, Tissue Expression, Literature, and Data) are available for data downloading and browsing, functional gene exploration, and experimental practice. A community forum is also available for information sharing, and a summary of current research progress on different aspects of lettuce is included. We believe that LettuceGDB can be a comprehensive functional database amenable to data mining and database-driven exploration, useful for both scientific research and lettuce breeding.

High-Throughput Phenotyping Accelerates the Dissection of the Phenotypic Variation and Genetic Architecture of Shank Vascular Bundles in Maize (Zea mays L.).

The vascular bundle of the shank is an important 'flow' organ for transforming maize biological yield to grain yield, and its microscopic phenotypic characteristics and genetic analysis are of great significance for promoting the breeding of new varieties with high yield and good quality. In this study, shank CT images were obtained using the standard process for stem micro-CT data acquisition at resolutions up to 13.5 µm. Moreover, five categories and 36 phenotypic traits of the shank including related to the cross-section, epidermis zone, periphery zone, inner zone and vascular bundle were analyzed through an automatic CT image process pipeline based on the functional zones. Next, we analyzed the phenotypic variations in vascular bundles at the base of the shank among a group of 202 inbred lines based on comprehensive phenotypic information for two environments. It was found that the number of vascular bundles in the inner zone (IZ_VB_N) and the area of the inner zone (IZ_A) varied the most among the different subgroups. Combined with genome-wide association studies (GWAS), 806 significant single nucleotide polymorphisms (SNPs) were identified, and 1245 unique candidate genes for 30 key traits were detected, including the total area of vascular bundles (VB_A), the total number of vascular bundles (VB_N), the density of the vascular bundles (VB_D), etc. These candidate genes encode proteins involved in lignin, cellulose synthesis, transcription factors, material transportation and plant development. The results presented here will improve the understanding of the phenotypic traits of maize shank and provide an important phenotypic basis for high-throughput identification of vascular bundle functional genes of maize shank and promoting the breeding of new varieties with high yield and good quality.

Quantitative phenotyping and evaluation for lettuce leaves of multiple semantic components.

BACKGROUND: Classification and phenotype identification of lettuce leaves urgently require fine quantification of their multi-semantic traits. Different components of lettuce leaves undertake specific physiological functions and can be quantitatively described and interpreted using their observable properties. In particular, petiole and veins determine mechanical support and material transport performance of leaves, while other components may be closely related to photosynthesis. Currently, lettuce leaf phenotyping does not accurately differentiate leaf components, and there is no comparative evaluation for positive-back of the same lettuce leaf. In addition, a few traits of leaf components can be measured manually, but it is time-consuming, laborious, and inaccurate. Although several studies have been on image-based phenotyping of leaves, there is still a lack of robust methods to extract and validate multi-semantic traits of large-scale lettuce leaves automatically. RESULTS: In this study, we developed an automated phenotyping pipeline to recognize the components of detached lettuce leaves and calculate multi-semantic traits for phenotype identification. Six semantic segmentation models were constructed to extract leaf components from visible images of lettuce leaves. And then, the leaf normalization technique was used to rotate and scale different leaf sizes to the "size-free" space for consistent leaf phenotyping. A novel lamina-based approach was also utilized to determine the petiole, first-order vein, and second-order veins. The proposed pipeline contributed 30 geometry-, 20 venation-, and 216 color-based traits to characterize each lettuce leaf. Eleven manually measured traits were evaluated and demonstrated high correlations with computation results. Further, positive-back images of leaves were used to verify the accuracy of the proposed method and evaluate the trait differences. CONCLUSIONS: The proposed method lays an effective strategy for quantitative analysis of detached lettuce leaves' fine structure and components. Geometry, color, and vein traits of lettuce leaf and its components can be comprehensively utilized for phenotype identification and breeding of lettuce. This study provides valuable perspectives for developing automated high-throughput phenotyping application of lettuce leaves and the improvement of agronomic traits such as effective photosynthetic area and vein configuration.

Dissecting the Genetic Structure of Maize Leaf Sheaths at Seedling Stage by Image-Based High-Throughput Phenotypic Acquisition and Characterization.

The rapid development of high-throughput phenotypic detection techniques makes it possible to obtain a large number of crop phenotypic information quickly, efficiently, and accurately. Among them, image-based phenotypic acquisition method has been widely used in crop phenotypic identification and characteristic research due to its characteristics of automation, non-invasive, non-destructive and high throughput. In this study, we proposed a method to define and analyze the traits related to leaf sheaths including morphology-related, color-related and biomass-related traits at V6 stage. Next, we analyzed the phenotypic variation of leaf sheaths of 418 maize inbred lines based on 87 leaf sheath-related phenotypic traits. In order to further analyze the mechanism of leaf sheath phenotype formation, 25 key traits (2 biomass-related, 19 morphology-related and 4 color-related traits) with heritability greater than 0.3 were analyzed by genome-wide association studies (GWAS). And 1816 candidate genes of 17 whole plant leaf sheath traits and 1,297 candidate genes of 8 sixth leaf sheath traits were obtained, respectively. Among them, 46 genes with clear functional descriptions were annotated by single nucleotide polymorphism (SNPs) that both Top1 and multi-method validated. Functional enrichment analysis results showed that candidate genes of leaf sheath traits were enriched into multiple pathways related to cellular component assembly and organization, cell proliferation and epidermal cell differentiation, and response to hunger, nutrition and extracellular stimulation. The results presented here are helpful to further understand phenotypic traits of maize leaf sheath and provide a reference for revealing the genetic mechanism of maize leaf sheath phenotype formation.