Comparative transcriptome analysis and genetic dissection of vegetative branching traits in foxtail millet (Setaria italica).

KEY MESSAGE: Two major genetic loci, qTN5.1 and qAB9.1, were identified and finely mapped to the 255 Kb region with one potential candidate gene for tiller number and the 521 Kb region with eight candidate genes for axillary branch number, respectively. Vegetative branching including tillering and axillary branching are vital traits affecting both the plant architecture and the biomass in cereal crops. However, the mechanism underlying the formation of vegetative branching in foxtail millet is largely unknown. Here, a foxtail millet cultivar and its bushy wild relative Setaria viridis accession were used to construct segregating populations to identify candidate genes regulating tiller number and axillary branch number. Transcriptome analysis using vegetative branching bud samples of parental accessions was performed, and key differentially expressed genes and pathways regulating vegetative branching were pointed out. Bulk segregant analysis on their F2:3 segregating population was carried out, and a major QTL for tiller number (qTN5.1) and two major QTLs for axillary branch number (qAB2.1 and qAB9.1) were detected. Fine-mapping strategy was further performed on F2:4 segregate population, and Seita.5G356600 encoding ß-glucosidase 11 was identified as the promising candidate gene for qTN5.1, and eight genes, especially Seita.9G125300 and Seita.9G125400 annotated as B-S glucosidase 44, were finally identified as candidate genes for regulating axillary branching. Findings in this study will help to elucidate the genetic basis of the vegetative branching formation of foxtail millet and lay a foundation for breeding foxtail millet varieties with ideal vegetative branching numbers.

Genome-wide identification of GRF transcription factors and their expression profile in stem meristem of broomcorn millet (Panicum miliaceum L.).

To understand the genome-wide information of the GRF family genes in broomcorn millet and their expression profile in the vegetative meristems, bioinformatic methods and transcriptome sequencing were used to analyze the characteristics, physical and chemical properties, phylogenetic relationship, chromosome distribution, gene structure, cis-acting elements and expression profile in stem meristem for the GRF family members. The results showed that the GRF gene family of millet contains 21 members, and the PmGRF gene is unevenly distributed on 12 chromosomes. The lengths of PmGRF proteins vary from 224 to 618 amino acids, and the isoelectric points are between 4.93-9.69. Each member of the family has 1-4 introns and 2-5 exons. The protein PmGRF13 is localized in both the nucleus and chloroplast, and the rest PmGRF proteins are located in the nucleus. Phylogenetic analysis showed that the 21 GRF genes were divided into 4 subfamilies (A,B,C and D) in broomcorn millet. The analysis of cis-acting elements showed that there were many cis-acting elements involved in light response, hormone response, drought induction, low temperature response and other environmental stress responses in the 2000 bp sequence upstream of the GRF genes. Transcriptome sequencing and qRT-PCR analyses showed that the expression levels of PmGRF3 and PmGRF12 in the dwarf variety Zhang778 were significantly higher than those of the tall variety Longmi12 in the internode and node meristems at the jointing stage, while the expression patterns of PmGRF4, PmGRF16 and PmGRF21 were reverse. In addition, the expression levels of PmGRF2 and PmGRF5 in the internode of Zhang778 were significantly higher than Longmi12. The other GRF genes were not or insignificantly expressed. These results indicated that seven genes, PmGRF2, PmGRF3, PmGRF4, PmGRF5, PmGRF12, PmGRF16 and PmGRF21, were related to the formation of plant height in broomcorn millet.

Genome-wide identification of quantitative trait loci for morpho-agronomic and yield-related traits in foxtail millet (Setaria italica) across multi-environments.

Foxtail millet (Setaria italica) is an ideal model of genetic system for functional genomics of the Panicoideae crop. Identification of QTL responsible for morpho-agronomic and yield-related traits facilitates dissection of genetic control and breeding in cereal crops. Here, based on a Yugu1 × Longgu7 RIL population and genome-wide resequencing data, an updated linkage map harboring 2297 bin and 74 SSR markers was constructed, spanning 1315.1 cM with an average distance of 0.56 cM between adjacent markers. A total of 221 QTL for 17 morpho-agronomic and yield-related traits explaining 5.5 ~ 36% of phenotypic variation were identified across multi-environments. Of these, 109 QTL were detected in two to nine environments, including the most stable qLMS6.1 harboring a promising candidate gene Seita.6G250500, of which 70 were repeatedly identified in different trials in the same geographic location, suggesting that foxtail millet has more identical genetic modules under the similar ecological environment. One hundred-thirty QTL with overlapping intervals formed 22 QTL clusters. Furthermore, six superior recombinant inbred lines, RIL35, RIL48, RIL77, RIL80, RIL115 and RIL125 with transgressive inheritance and enrichment of favorable alleles in plant height, tiller, panicle morphology and yield related-traits were screened by hierarchical cluster. These identified QTL, QTL clusters and superior lines lay ground for further gene-trait association studies and breeding practice in foxtail millet.

QTL mapping of yield component traits on bin map generated from resequencing a RIL population of foxtail millet (Setaria italica).

BACKGROUND: Foxtail millet (Setaria italica) has been developed into a model genetical system for deciphering architectural evolution, C4 photosynthesis, nutritional properties, abiotic tolerance and bioenergy in cereal grasses because of its advantageous characters with the small genome size, self-fertilization, short growing cycle, small growth stature, efficient genetic transformation and abundant diverse germplasm resources. Therefore, excavating QTLs of yield component traits, which are closely related to aspects mentioned above, will further facilitate genetic research in foxtail millet and close cereal species. RESULTS: Here, 164 Recombinant inbreed lines from a cross between Longgu7 and Yugu1 were created and 1,047,978 SNPs were identified between both parents via resequencing. A total of 3413 bin markers developed from SNPs were used to construct a binary map, containing 3963 recombinant breakpoints and totaling 1222.26 cM with an average distance of 0.36 cM between adjacent markers. Forty-seven QTLs were identified for four traits of straw weight, panicle weight, grain weight per plant and 1000-grain weight. These QTLs explained 5.5-14.7% of phenotypic variance. Thirty-nine favorable QTL alleles were found to inherit from Yugu1. Three stable QTLs were detected in multi-environments, and nine QTL clusters were identified on Chromosome 3, 6, 7 and 9. CONCLUSIONS: A high-density genetic map with 3413 bin markers was constructed and three stable QTLs and 9 QTL clusters for yield component traits were identified. The results laid a powerful foundation for fine mapping, identifying candidate genes, elaborating molecular mechanisms and application in foxtail millet breeding programs by marker-assisted selection.

A high density genetic map and QTL for agronomic and yield traits in Foxtail millet [Setaria italica (L.) P. Beauv].

BACKGROUND: Foxtail millet [Setaria italica (L.) P. Beauv.], a crop of historical importance in China, has been adopted as a model crop for studying C-4 photosynthesis, stress biology and biofuel traits. Construction of a high density genetic map and identification of stable quantitative trait loci (QTL) lay the foundation for marker-assisted selection for agronomic traits and yield improvement. RESULT: A total of 10598 SSR markers were developed according to the reference genome sequence of foxtail millet cultivar 'Yugu1'. A total of 1013 SSR markers showing polymorphism between Yugu1 and Longgu7 were used to genotype 167 individuals from a Yugu1 × Longgu7 F2 population, and a high density genetic map was constructed. The genetic map contained 1035 loci and spanned 1318.8 cM with an average distance of 1.27 cM between adjacent markers. Based on agronomic and yield traits identified in 2 years, 29 QTL were identified for 11 traits with combined analysis and single environment analysis. These QTL explained from 7.0 to 14.3 % of phenotypic variation. Favorable QTL alleles for peduncle length originated from Longgu7 whereas favorable alleles for the other traits originated from Yugu1 except for qLMS6.1. CONCLUSIONS: New SSR markers, a high density genetic map and QTL identified for agronomic and yield traits lay the ground work for functional gene mapping, map-based cloning and marker-assisted selection in foxtail millet.

Phosphorus and naphthalene acetic acid increased the seed yield by regulating carbon and nitrogen assimilation of flax.

To evaluate the impact of phosphorus (P) combined with exogenous NAA on flax yield, enhance flax P utilization efficiency and productivity, minimize resource inputs and mitigate negative environmental and human effects. Therefore, it is crucial to comprehend the physiological and biochemical responses of flax to P and naphthylacetic acid (NAA) in order to guide future agronomic management strategies for increasing seed yield. A randomized complete block design trial was conducted under semi-arid conditions in Northwest China, using a factorial split-plot to investigate the effects of three P (0, 67.5, and 135.0 kg P2O5 ha-1) and three exogenous spray NAA levels (0, 20, and 40 mg NAA L-1) on sucrose phosphate synthase (SPS) and diphosphoribulose carboxylase (Rubisco) activities as well as nitrogen (N) and P accumulation and translocation in flax. Results indicated that the SPS and Rubisco activities, N and P accumulation at flowering and maturity along with assimilation and translocation post-flowering, fruiting branches per plant, tillers per plant, capsules per plant, and seed yield were 95, 105, 14, 27, 55, 15, 13, 110, 103, 82, 16, 61, 8, and 13% greater in the P treatments compared to those in the zero P treatment, respectively. Moreover, those characteristics were observed to be greater with exogenous spray NAA treatments compared to that no spray NAA treatment. Additionally, the maximum SPS and Rubisco activities, N and P accumulation, assimilation post-flowering and translocation, capsules per plant, and seed yield were achieved with the application of 67.5 kg P2O5 ha-1 with 20 mg NAA L-1. Therefore, these findings demonstrate that the appropriate combination of P fertilizer and spray NAA is an effective agronomic management strategy for regulating carbon and nitrogen assimilation by maintaining photosynthetic efficiency in plants to increase flax productivity.

Pangenome analysis reveals genomic variations associated with domestication traits in broomcorn millet.

Broomcorn millet (Panicum miliaceum L.) is an orphan crop with the potential to improve cereal production and quality, and ensure food security. Here we present the genetic variations, population structure and diversity of a diverse worldwide collection of 516 broomcorn millet genomes. Population analysis indicated that the domesticated broomcorn millet originated from its wild progenitor in China. We then constructed a graph-based pangenome of broomcorn millet based on long-read de novo genome assemblies of 32 representative accessions. Our analysis revealed that the structural variations were highly associated with transposable elements, which influenced gene expression when located in the coding or regulatory regions. We also identified 139 loci associated with 31 key domestication and agronomic traits, including candidate genes and superior haplotypes, such as LG1, for panicle architecture. Thus, the study's findings provide foundational resources for developing genomics-assisted breeding programs in broomcorn millet.

Identification and fine-mapping of a major QTL (PH1.1) conferring plant height in broomcorn millet (Panicum miliaceum).

The plant height of broomcorn millet (Panicum miliaceum) is a significant agronomic trait that is closely related to its plant architecture, lodging resistance, and final yield. However, the genes underlying the regulation of plant height in broomcorn millet are rarely reported. Here, an F2 population derived from a cross between a normal variety, "Longmi12," and a dwarf mutant, "Zhang778," was constructed. Genetic analysis for the F2 and F2:3 populations revealed that the plant height was controlled by more than one locus. A major quantitative trait locus (QTL), PH1.1, was preliminarily identified in chromosome 1 using bulked segregant analysis sequencing (BSA-seq). PH1.1 was fine-mapped to a 109-kb genomic region with 15 genes using a high-density map. Among them, longmi011482 and longmi011489, containing nonsynonymous variations in their coding regions, and longmi011496, covering multiple insertion/deletion sequences in the promoter regions, may be possible candidate genes for PH1.1. Three diagnostic markers closely linked to PH1.1 were developed to validate the PH1.1 region in broomcorn millet germplasm. These findings laid the foundation for further understanding of the molecular mechanism of plant height regulation in broomcorn millet and are also beneficial to the breeding program for developing new varieties with optimal height.

Genetic Diversity and Classification of the Cytoplasm of Chinese Elite Foxtail Millet [Setaria italica (L.) P. Beauv.] Parental Lines Revealed by Chloroplast Deoxyribonucleic Acid Variation.

Due to the maternal inheritance of cytoplasm, using foxtail millet [Setaria italica (L.) P. Beauv.] male sterile lines with a single cytoplasmic source as the female parent will inevitably lead to a narrow source of cytoplasm in hybrids, which may make them vulnerable to infection by cytoplasm-specific pathogens, ultimately leading to destructive yield losses. To assess cytoplasmic genetic diversity in plants, molecular markers derived from chloroplast DNA (cpDNA) have been used. However, such markers have not yet been applied to foxtail millet. In this study, we designed and screened nine pairs of polymorphic foxtail millet-specific primers based on its completely sequenced cpDNA. Using these primers, we analyzed the genetic diversity and cytoplasmic types of 130 elite foxtail millet parental lines collected in China. Our results revealed that the cytoplasmic genetic diversity of these accessions was low and needs to be increased. The parental lines were divided into four cytoplasmic types according to population structure analysis and a female parent-derivative evolutionary graph, indicating that the cytoplasmic types of elite foxtail millet lines were rather limited. A principal component analysis (PCA) plot was linked with the geographic and ecological distribution of accessions for each cytoplasmic type, as well as their basal maternal parents. Collectively, our results suggest that enriching cytoplasmic sources through the use of accessions from diverse ecological regions and other countries as the female parent may improve foxtail millet breeding programs, and prevent infection by cytoplasm-specific pathogens.