Bioscience Methods 2024, Vol.15, No.4, 149-161 http://bioscipublisher.com/index.php/bm 151 by the time of day, with peak expression of C4 cycle genes occurring between 6 and 10 hours after dawn, coinciding with maximum photosynthetic rates (Borba et al., 2023). Figure 1 Single-Cell RNA sequencing analysis of maize leaf cells (Adapted from Tao et al., 2022) Image caption: (A) UMAP-based scatterplot displaying eight distinct cell clusters identified from 7,354 filtered maize leaf cells. (B) Dot plot illustrating the average expression levels of 16 genes specific to various cell types, differentially expressed between mesophyll (M) cells, bundle sheath (BS) cells, and other cell types across the eight Seurat clusters. The dot size indicates the proportion of cells expressing each gene, with genes more highly expressed in BS cells highlighted on a grey background. (C) Scatter plot comparing log2 (mean RPM+1) values for all genes in bulk RNA-seq data (x-axis) and scRNA-seq data (y-axis), with each dot's color representing the log2-transformed number of cells expressing the gene in the scRNA-seq. The correlation between the two datasets was assessed using the Spearman correlation test (Adapted from Tao et al., 2022) 3.2 Regulation by light and circadian rhythms Light and circadian rhythms are critical regulators of photosynthesis in maize. The expression of photosynthesis-related genes is tightly regulated by light intensity and circadian rhythms, with maximal photosynthetic activity observed around midday (Borba et al., 2023). Light exposure significantly inhibits the elongation of maize mesocotyl and coleoptile by modulating phytohormone levels and lignin deposition, which in turn affects the expression of genes involved in circadian rhythm and phytohormone biosynthesis (Zhao et al., 2023). 3.3 Role of transcription factors and gene networks Transcription factors (TFs) play pivotal roles in regulating photosynthesis in maize. For instance, basic Helix-Loop-Helix (bHLH) TFs, such as ZmbHLH128 and ZmbHLH129, bind to the promoter of the NADP-Malic Enzyme (NADP-ME) gene, a key enzyme in C4 photosynthesis, indicating the importance of TFs in the regulation
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