**6. Potential Role of Bioinformatics in the Prediction of miRNA and cDNA Microarray**

Next-generation sequencing methods are crucial in gene expression profiling, epigenomics, genomics, and transcriptomics. These tools can sequence multiple DNA molecules within a short period. The recent introduction of innovative "-omics" technologies, such as metabolomics, proteomics, and genomics allows for analyzing and identifying the genetic elements that contribute to system complexity [72,90,174,175]. Bioinformatics tools developed for miRNA prediction include miRNA target prediction, analysis, and structure prediction. For example, miRanda, RNAhybrid, RNA22, and TarHunter detect miRNA expression and perform analysis based on miRNA-Seq data (Table 4). Existing plant miRNA prediction tools lack a cross-species conservation filter and eTM prediction function. TarHunter features a strict cross-species conservation filter and the capability of predicting eTMs [158]. Despite ongoing progress, bioinformatics prediction of microRNA targets remains difficult, since current tools have a lack of accuracy and sensitivity. [72,176]. Microarrays are an effective method for determining the quantity of RNA in a sample. Since microarray data have computational complexity and contain hundreds of genes, statistical and bioinformatics methods are required for data interpretation [160]. These specialized tools provide statistical analysis, sample comparisons, and functional interpretation of data generated in a series following visualization and normalization in a microarray study, such as Array Designer, eArray, Visual OMP, caArray, and dCHIP (Table 4). The software, including Able Image Analyser, Gene pix pro-6.0, and GeneChip operating software, are used for analyzing images in order to obtain the intensity at each spot and quantify the expression for each transcript. Additionally, this also provides different types of discoveries by comparing gene expression data with already reported biological information, such as protein–protein interactions, pathway analysis, transcription factor binding sites, and network analysis tools, including Array Designer, eArray, Significance Analysis of Microarrays, Gene Expression, and Model Selector (Table 4) [164].

### **7. Conclusions and Future Perspectives**

MicroRNAs (miRNAs) have been considered a potential target in genetic engineering against abiotic stresses in plants. Thus, miRNAs can also be utilized in the initial monitoring and transmission of abiotic stresses, and to elucidate the genetic and physiological responses against stress in plants. This review summarized current developments and the history of miRNAs and microarray with diverse functions in several stress responses, predominantly abiotic stresses. Many traditional approaches have identified significant numbers of miRNAs in plants from various organisms. Microarray-based genomic technologies for ecological studies have received great attention, particularly in plants, to disclose the role of stress-responsive loci in plants. DNA microarrays provide a novel insight into the cell and provide a solution for several problems from the viewpoint of analytical calculation, despite the inconceivable amount of work done in

the last two decades to reduce the different sources of uncertainty on the subsequent measurements. The review will provide valuable insight to plant researchers, especially plant breeders and stress physiologists, to design a comprehensive strategy to cope with environmental stresses.

The elucidation of miRNA responses to abiotic stresses may lead to the development of technologies for the early detection of plants' environmental stressors. MicroRNAs and cDNA-microarrays are powerful targets for engineering abiotic stress tolerance in transgenic plants. The field of bioinformatics is developing rapidly, and it is inevitable to progress in plant genomics and breeding without integrating the latest bioinformatics tools. Multiple advanced sequencing and bioinformatics tools were established to identify miRNAs and their target gene network and prediction. As the understanding of the function of miRNAs under stress deepens, the potential use of miRNA mediated genes to enhance plant tolerance will also increase. In the future, the large-scale microarrays might be substituted with small biosensors which contain a unique or a small number of novel microbes deposited on an electronic platform. We would like to conclude by illustrating the existing gap between the detection of stress-regulated miRNAs and microarray to validate their role. In conclusion, we recommend the utilization of miRNAs for the identification and classification of new functional genes conferring a significant functional role in stress tolerance and to exploit the unexplained fraction of genes further.

**Author Contributions:** T.P. and M.W.A.: have made substantial contributions to formulate, Drafting and retrieve data, A.E.-k.; revised and improved the technical language S.-H.N.; Revised and improved the MS, H.X.W.; Final approval and revised for publication. All authors have read and agreed to the published version of the manuscript.

**Funding:** This work is funded by the National Natural Science Foundation of China (31870651).

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** Not applicable.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**

