Transcriptome Sequencing and Metabolome Analysis Reveals the Molecular Mechanism of Drought Stress in Millet.

As one of the oldest agricultural crops in China, millet (Panicum miliaceum) has powerful drought tolerance. In this study, transcriptome and metabolome analyses of 'Hequ Red millet' (HQ) and 'Yanshu No.10' (YS10) millet after 6 h of drought stress were performed. Transcriptome characteristics of drought stress in HQ and YS10 were characterized by Pacbio full-length transcriptome sequencing. The pathway analysis of the differentially expressed genes (DEGs) showed that the highly enriched categories were related to starch and sucrose metabolism, pyruvate metabolism, metabolic pathways, and the biosynthesis of secondary metabolites when the two millet varieties were subjected to drought stress. Under drought stress, 245 genes related to energy metabolism were found to show significant changes between the two strains. Further analysis showed that 219 genes related to plant hormone signal transduction also participated in the drought response. In addition, numerous genes involved in anthocyanin metabolism and photosynthesis were confirmed to be related to drought stress, and these genes showed significant differential expression and played an important role in anthocyanin metabolism and photosynthesis. Moreover, we identified 496 transcription factors related to drought stress, which came from 10 different transcription factor families, such as bHLH, C3H, MYB, and WRKY. Further analysis showed that many key genes related to energy metabolism, such as citrate synthase, isocitrate dehydrogenase, and ATP synthase, showed significant upregulation, and most of the structural genes involved in anthocyanin biosynthesis also showed significant upregulation in both strains. Most genes related to plant hormone signal transduction showed upregulated expression, while many JA and SA signaling pathway-related genes were downregulated. Metabolome analysis was performed on 'Hequ red millet' (HQ) and 'Yanshu 10' (YS10), a total of 2082 differential metabolites (DEMs) were identified. These findings indicate that energy metabolism, anthocyanins, photosynthesis, and plant hormones are closely related to the drought resistance of millet and adapt to adversity by precisely regulating the levels of various molecular pathways.

Application of protoplast technology to CRISPR/Cas9 mutagenesis: from single-cell mutation detection to mutant plant regeneration.

Plant protoplasts are useful for assessing the efficiency of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) mutagenesis. We improved the process of protoplast isolation and transfection of several plant species. We also developed a method to isolate and regenerate single mutagenized Nicotianna tabacum protoplasts into mature plants. Following transfection of protoplasts with constructs encoding Cas9 and sgRNAs, target gene DNA could be amplified for further analysis to determine mutagenesis efficiency. We investigated N. tabacum protoplasts and derived regenerated plants for targeted mutagenesis of the phytoene desaturase (NtPDS) gene. Genotyping of albino regenerants indicated that all four NtPDS alleles were mutated in amphidiploid tobacco, and no Cas9 DNA could be detected in most regenerated plants.

Exploration of millet models for developing nutrient rich graminaceous crops.

Protein-energy malnutrition and micronutrient deficiencies contribute to high mortality among considerable proportion of the current 7.2 billion global populations, especially children. Although poverty and diets poor in nutrition are prime reasons for prevalence of malnutrition, nutritionally dense crops offer an inexpensive and sustainable solution to the problem of malnutrition. Remarkably, millets are nutritionally superior to major non-millet cereals. They especially are rich in dietary fibers, antioxidants, phytochemicals and polyphenols, which contribute broad-spectrum positive impacts to human health. However, millets have received lesser research attention universally, and considering this, the present review was planned to summarize the reports available on nutrition profile of millets and non-millet cereals to provide a comparative insight on importance of millets. It also emphasizes the need for research on deciphering nutritional traits present in millets and to develop strategies for introgressing these traits into other conventional staple crops using germplasm and 'omics' technologies. In some millet species, excellent 'omics' and germplasm panels have started to get available which can act as a starting point for understanding as well as of introgressing healthful traits across millets and non-millet cereals.