Cotton Genomics and Genetics 2025, Vol.16, No.4, 184-191 http://cropscipublisher.com/index.php/cgg 185 2 Applications of Transcriptomics in Plant Immunity Research 2.1 RNA-Seq technology and its advantages RNA-Seq is a technology used to analyze RNA expression. It can measure the types and quantities of all RNA in a sample at one time. This technology is more sensitive than the older generation of methods (such as chips), can find new gene transcripts, and can more accurately calculate the expression level of each gene. When studying plant immune responses, RNA-Seq can help us understand which genes are activated and when, and which tissues are changing. This can reveal the details of plant immune changes, such as discovering immune channels or regulatory genes that were not noticed before, such as calcium channels or specific transcription factors (Moore et al., 2011; Bjornson et al., 2020; Zheng et al., 2024). 2.2 Cotton transcriptome databases and resource integration Integrating transcriptome data from different experiments is the key to studying plant immunity. There are many public databases that collect RNA data from different plants and different experimental conditions, which can be used for comparative analysis. These data help scientists find some common immune-related genes, such as antimicrobial peptides. With these resources, researchers can also develop some computational tools to discover new genes or regulatory sites (Shelenkov et al., 2020; De Jong and Bosco, 2021). 2.3 Bioinformatics methods and analytical pipeline Bioinformatics plays a big role in analyzing RNA-Seq data. Generally speaking, the analysis process starts with a data quality check, then the reads are aligned to the genome, then which genes have changed in expression, and finally the functions of these genes are checked. Some new algorithms can find which genes work together, which regulatory modules are at work, and which pathways are involved in the immune response. This type of analysis helps us understand the activity of genes in different time and space, and can also predict some new immune-related genes or small peptides (Zhang et al., 2017; De Jong and Bosco, 2021; Liu et al., 2024). Now, researchers have also begun to combine batch sequencing and single-cell sequencing data, which can see more details and further understand the mechanisms behind plant immune responses (Stubbington et al., 2017). 3 Gene Expression Changes in Cotton under Biotic Stress 3.1 Transcriptional regulation under pathogen stress When cotton encounters pathogen invasion, a complex transcriptional response mechanism is quickly activated. Transcription factors such as WRKY28, WRKY40, and WRKY53, as well as proteins such as ERF and bHLH, will rise rapidly in leaves. They can activate hormone signals (such as jasmonic acid, salicylic acid, and ethylene) and regulate the synthesis of some secondary metabolites to help cotton resist pathogens. Studies have found that transcription factors of the WRKY and ERF families can be activated under a variety of biotic stresses, indicating that they play a core role in defense (Liu et al., 2016; Bihani et al., 2024; Huang et al., 2024). There is also a gene called GhJAZ2, which can also regulate the jasmonic acid pathway, helping cotton find a balance in defense between pathogens and leaf-eating insects (Figure 1) (He et al., 2018). 3.2 Differential gene expression induced by insect infestation Insects such as aphids, cotton bollworms, and whiteflies can cause a series of genetic changes in cotton. Some transcription factors, such as WRKY28, WRKY40, WRKY53, ERF4 and ERF5, are activated when different insects invade, which is a more "universal" response. But there are also some genes that are "specific for a certain insect", such as GH3.1, ACS1, CYP74A, TIFY10A, BHLH25, ABR1, ERF025, which are particularly active when cotton bollworm invades. These genes are responsible for regulating hormone responses, MAPK signaling pathways and the production of some defensive metabolites, allowing cotton to respond more accurately to different insects (Bihani et al., 2024). 3.3 Integration of immune pathways under multiple stresses Sometimes, cotton may face attacks from pathogens and insects at the same time. At this time, it will activate multiple defense channels to work together. Some genes, such as GhWRKY28, are activated when encountering abiotic stresses such as diseases and drought, indicating that these defense signals can be combined with each
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