Development and application of whole-chromosome painting of chromosomes 7A and 8A of Arachis duranensis based on chromosome-specific single-copy oligonucleotides.

For peanut, the lack of stable cytological markers is a barrier to tracking specific chromosomes, elucidating the genetic relationships between genomes and identifying chromosomal variations. Chromosome mapping using single-copy oligonucleotide (oligo) probe libraries has unique advantages for identifying homologous chromosomes and chromosomal rearrangements. In this study, we developed two whole-chromosome single-copy oligo probe libraries, LS-7A and LS-8A, based on the reference genome sequences of chromosomes 7A and 8A of Arachis duranensis. Fluorescence in situ hybridization (FISH) analysis confirmed that the libraries could specifically paint chromosomes 7 and 8. In addition, sequential FISH and electronic localization of LS-7A and LS-8A in A. duranensis (AA) and A. ipaensis (BB) showed that chromosomes 7A and 8A contained translocations and inversions relative to chromosomes 7B and 8B. Analysis of the chromosomes of wild Arachis species using LS-8A confirmed that this library could accurately and effectively identify A genome species. Finally, LS-7A and LS-8A were used to paint the chromosomes of interspecific hybrids and their progenies, which verified the authenticity of the interspecific hybrids and identified a disomic addition line. This study provides a model for developing specific oligo probes to identify the structural variations of other chromosomes in Arachis and demonstrates the practical utility of LS-7A and LS-8A.

Transcriptome analysis of pod mutant reveals plant hormones are important regulators in controlling pod size in peanut (Arachis hypogaea L.).

Pod size is an important yield-influencing trait in peanuts. It is affected by plant hormones and identifying the genes related to these hormones may contribute to pod-related trait improvements in peanut breeding programs. However, there is limited information on the molecular mechanisms of plant hormones that regulate pod size in peanuts. We identified a mutant with an extremely small pod (spm) from Yuanza 9102 (WT) by 60Co γ-radiation mutagenesis. The length and width of the natural mature pod in spm were only 71.34% and 73.36% of those in WT, respectively. We performed comparative analyses for morphological characteristics, anatomy, physiology, and global transcriptome between spm and WT pods. Samples were collected at 10, 20, and 30 days after peg elongation into the soil, representing stages S1, S2, and S3, respectively. The differences in pod size between WT and spm were seen at stage S1 and became even more striking at stages S2 and S3. The cell sizes of the pods were significantly smaller in spm than in WT at stages S1, S2, and S3. These results suggested that reduced cell size may be one of the important contributors for the small pod in spm. The contents of indole-3-acetic acid (IAA), gibberellin (GA), and brassinosteroid (BR) were also significantly lower in spm pods than those in WT pods at all three stages. RNA-Seq analyses showed that 1,373, 8,053, and 3,358 differently expressed genes (DEGs) were identified at stages S1, S2, and S3, respectively. Functional analyses revealed that a set of DEGs was related to plant hormone biosynthesis, plant hormone signal transduction pathway, and cell wall biosynthesis and metabolism. Furthermore, several hub genes associated with plant hormone biosynthesis and signal transduction pathways were identified through weighted gene co-expression network analysis. Our results revealed that IAA, GA, and BR may be important regulators in controlling pod size by regulating cell size in peanuts. This study provides helpful information for the understanding of the complex mechanisms of plant hormones in controlling pod size by regulating the cell size in peanuts and will facilitate the improvement of peanut breeding.

Mapping of a QTL associated with sucrose content in peanut kernels using BSA-seq.

As an important factor affecting the edible quality of peanut kernels, sucrose content is a complex quantitative trait regulated by multiple factors. In this study, an F2 segregating population and a recombinant inbred line (RIL) population, derived from a cross between the high sucrose content variety Jihuatian 1 and the low sucrose content line PI478819, were used as materials to map a quantitative trait locus (QTL) associated with sucrose content in peanut kernels. Four QTLs were initially located on chromosomes A03 and A06 based on BSA-seq technology, and multiple kompetitive allele-specific PCR markers were developed based on single-nucleotide polymorphisms (SNPs) in the intervals. The markers were genotyped in the RIL population and finely mapped to a stable QTL, qSUCA06, located on chromosome A06 within a 0.29-Mb physical genomic interval (112367085-112662675 bp), which accounted for 31.95%-41.05% of the phenotypic variance explained. SNP and insertion/deletion annotations were performed on genes in the candidate interval, and having screened out those genes with mutations in exons, candidate genes were verified by qRT-PCR. The results revealed that Arahy.Y2LWD9 may be the main gene regulating sucrose content. The QTL identified in this study will not only contribute to marker-assisted breeding for improvement of peanut sucrose content but also paves the way for identifying gene function.

Third-generation sequencing and metabolome analysis reveal candidate genes and metabolites with altered levels in albino jackfruit seedlings.

BACKGROUND: Most plants rely on photosynthesis; therefore, albinism in plants with leaves that are white instead of green causes slow growth, dwarfing, and even death. Although albinism has been characterized in annual model plants, little is known about albino trees. Jackfruit (Artocarpus heterophyllus) is an important tropical fruit tree species. To gain insight into the mechanisms underlying the differential growth and development between albino jackfruit mutants and green seedlings, we analyzed root, stem, and leaf tissues by combining PacBio single-molecule real-time (SMRT) sequencing, high-throughput RNA-sequencing (RNA-seq), and metabolomic analysis. RESULTS: We identified 8,202 differentially expressed genes (DEGs), including 225 genes encoding transcription factors (TFs), from 82,572 full-length transcripts. We also identified 298 significantly changed metabolites (SCMs) in albino A. heterophyllus seedlings from a set of 692 metabolites in A. heterophyllus seedlings. Pathway analysis revealed that these DEGs were highly enriched in metabolic pathways such as 'photosynthesis', 'carbon fixation in photosynthetic organisms', 'glycolysis/gluconeogenesis', and 'TCA cycle'. Analysis of the metabolites revealed 76 SCMs associated with metabolic pathways in the albino mutants, including L-aspartic acid, citric acid, succinic acid, and fumaric acid. We selected 225 differentially expressed TF genes, 333 differentially expressed metabolic pathway genes, and 76 SCMs to construct two correlation networks. Analysis of the TF-DEG network suggested that basic helix-loop-helix (bHLH) and MYB-related TFs regulate the expression of genes involved in carbon fixation and energy metabolism to affect light responses or photomorphogenesis and normal growth. Further analysis of the DEG-SCM correlation network and the photosynthetic carbon fixation pathway suggested that NAD-ME2 (encoding a malic enzyme) and L-aspartic acid jointly inhibit carbon fixation in the albino mutants, resulting in reduced photosynthetic efficiency and inhibited plant growth. CONCLUSIONS: Our preliminarily screening identified candidate genes and metabolites specifically affected in albino A. heterophyllus seedlings, laying the foundation for further study of the regulatory mechanism of carbon fixation during photosynthesis and energy metabolism. In addition, our findings elucidate the way genes and metabolites respond in albino trees.

Third-Generation Sequencing Reveals LncRNA-Regulated HSP Genes in the Populus x canadensis Moench Heat Stress Response.

Long non-coding RNAs (lncRNAs) regulate plant responses to abiotic stresses. However, the short reads produced by second-generation sequencing technology make it difficult to accurately explore full-length transcripts, limiting the study of lncRNAs. In this study, we used third-generation long-read sequencing technology with the PacBio Sequel and Illumina platform to explore the role of lncRNAs in the heat stress response of Populus x canadensis Moench trees. We using 382,034,416 short reads to correct 4,297,179 long reads by resulted in 66,657 full-length transcripts, representing 33,840 genes. Then, 753 putative lncRNAs were identified, including 658 sense lncRNAs (87.38%), 41 long intervening/intergenic non-coding RNAs (lincRNAs) (5.44%), 12 antisense lncRNAs (1.59%), and 42 sense intronic lncRNAs (5.58%). Using the criteria | log2FC| ≥ 1 and q-value < 0.05, 3,493 genes and 78 lncRNAs were differentially expressed under the heat treatment. Furthermore, 923 genes were detected as targets of 43 differently expressed lncRNAs by cis regulation. Functional annotation demonstrated that these target genes were related to unfolded protein binding, response to stress, protein folding, and response to stimulus. Lastly, we identified a lncRNA-gene interaction network consisting of four lncRNAs and six genes [Heat Shock Protein 82 (HSP82), HSP83, Disease Resistance Protein 27 (DRL27), DnaJ family protein (DNJH), and two other predicted protein-coding genes], which showed that lncRNAs could regulate HSP family genes in response to heat stress in Populus. Therefore, our third-generation sequencing has improved the description of the P. canadensis transcriptome. The potential lncRNAs and HSP family genes identified here present a genetic resource to improve our understanding of the heat-adaptation mechanisms of trees.