Assessing nutritional and genetic variations within foxtail millet (Setaria italica) landraces collected from indigenous communities across the Philippines.

Unknown to many, the Philippines is host to a few remaining accessions of the underutilised and understudied cereal foxtail millet (Setaria italica (L.) P. Beauv.). We collected together accessions of this crop from different eco-geographical locations within the Philippines, along with a few accessions from Lanyu, Taiwan, to undertake a study of their nutritional value and genetic diversity. All accessions were field-grown in 2022, dry season (DS) at the Institute of Plant Breeding (IPB) Experiment Station, Los Baños, Laguna, Philippines. The accessions were tested for micronutrients, including Zn and Fe, nitrogen as a proxy for protein, ß-carotene, phytic acid, and a number of phenolic compounds with known nutritional potential. Of the 20 accessions tested, the accessions Bayaras and GB61438 had the highest level of Zn (107.1 mg/kg) and Fe (70.52 mg/kg), respectively, higher than levels found in traditional rice varieties. For ß-carotene the highest concentration was found in the accession Balles (∼10 µg/g). Twelve phenolic compounds were detected, with catechin, syringic acid, ferulic acid and kaempferol having the highest concentrations. To assess the genetic diversity, we sequenced a set of eight samples selected from among the accessions to a depth of at least 25-fold using whole-genome re-sequencing. Analysis of the population structure, using genome-wide, high-quality SNPs, showed modest diversity among the accessions, with two unadmixed groups. The accessions are monophyletic relative to their earliest common ancestor, with the very light brown accessions emerging earlier than the light brown and reddish-brown varieties. Analysis of zinc-regulated, iron-regulated transporter-like protein (ZIP) transporters within the foxtail millet reference sequence, var. Yugu1 identified 17 putative ZIP transporters. Variant calling identified SNPs primarily within 3' and 5' regions, and introns, indicating variation between foxtail millet accessions within regulatory gene regions rather than in structural proteins. The local foxtail millet accessions, therefore, represent a potential alternative source of nutrients which may help in addressing malnutrition in the Philippines.

Assessing Loss of Regulatory Divergence, Genome-Transcriptome Incongruence, and Preferential Expression Switching in Abaca × Banana Backcrosses.

The Musa textilis var. Abuab has high fiber quality (FQ) but is susceptible to abaca bunchy top virus (AbBTV); the Musa balbisiana var. Pacol has low FQ but is resistant against AbBTV. Their backcrosses (BC2 and BC3) possess both desirable traits. Analysis using RNA-seq showed that the regulatory divergence of Abuab and Pacol is largely explained by cis differences with 27.4% and 22.3% if we are to assess it using BC2 and BC3, respectively. Cis differences between the two genotypes are significantly reduced from BC2 to BC3 due to changes in genomic constitution. Trans, on the other hand, is robust to changes in allelic composition. All these are attributed to the loss of heterozygosity in BC3 relative to BC2. Further analysis showed that both backcrosses exhibited genome-wide preferential expression of Pacol- over Abuab-specific alleles, despite the wider genetic presence of the latter in the hybrids. The ratio of the two genotype-specific expressed transcripts and the ratio of their corresponding genetic make-up are significantly disproportionate, a phenomenon that we refer to here as "genome-transcriptome incongruence". We also observed preferential expression switching in which several genes prefer the Abuab- (or Pacol-) specific allele in BC2 but switched to the Pacol- (or Abuab-) specific allele in the BC3 genome.

Nutritional and genetic variation in a core set of Ethiopian Tef (Eragrostis tef) varieties.

BACKGROUND: Tef (Eragrostis tef) is a tropical cereal domesticated and grown in the Ethiopian highlands, where it has been a staple food of Ethiopians for many centuries. Food insecurity and nutrient deficiencies are major problems in the country, so breeding for enhanced nutritional traits, such as Zn content, could help to alleviate problems with malnutrition. RESULTS: To understand the breeding potential of nutritional traits in tef a core set of 24 varieties were sequenced and their mineral content, levels of phytate and protein, as well as a number of nutritionally valuable phenolic compounds measured in grain. Significant variation in all these traits was found between varieties. Genome wide sequencing of the 24 tef varieties revealed 3,193,582 unique SNPs and 897,272 unique INDELs relative to the tef reference var. Dabbi. Sequence analysis of two key transporter families involved in the uptake and transport of Zn by the plant led to the identification of 32 Zinc Iron Permease (ZIP) transporters and 14 Heavy Metal Associated (HMA) transporters in tef. Further analysis identified numerous variants, of which 14.6% of EtZIP and 12.4% of EtHMA variants were non-synonymous changes. Analysis of a key enzyme in flavanol synthesis, flavonoid 3'-hydroxylase (F3'H), identified a T-G variant in the tef homologue Et_s3159-0.29-1.mrna1 that was associated with the differences observed in kaempferol glycoside and quercetin glycoside levels. CONCLUSION: Wide genetic and phenotypic variation was found in 24 Ethiopian tef varieties which would allow for breeding gains in many nutritional traits of importance to human health.

RNA-Seq Reveals Differentially Expressed Genes Associated with High Fiber Quality in Abaca (Musa textilis Nee).

Despite the importance of and current demand for abaca (Musa textilis Nee) fiber, there has been limited study that capitalizes on RNA-seq to identify candidate genes associated with high fiber quality and bunchy top virus (AbBTV) resistance. Three varieties (Abuab, Inosa, and Tangongon), one wild banana variety (Musa balbisiana Colla) Pacol, and two developed backcrosses (Abuab × Pacol BC2 and BC3) were grown at the Institute of Plant Breeding (IPB), Laguna, Philippines. The pseudostems of 3-month-old suckers of each genotype were sampled for RNA-seq. Datasets were analyzed for differential expression (DE) implementing various model frameworks, including pairwise, genotypic and non-DE models. Results indicate that Abuab and BC3 induce the highest proportion (70%) of abaca-specific genes. Gene ontology (GO) enrichment analysis showed several genes associated with cellulose synthase activity, callose synthase, ß-glucosidase activity, glucan biosynthetic process, etc. KEGG pathway analysis showed several genes encoding for enzymes involved in the lignin biosynthetic pathway. Analysis using genotypic DE (GDE) between abaca bunchy top virus (AbBTV)-resistant and -susceptible groups revealed genes such as pathogenesis-related protein and NBS-LRR. As the genotypes were not infected with the pathogen, these genes are yet to be confirmed for their roles in disease resistance and are an interesting subject for further investigation.

Unraveling regulatory divergence, heterotic malleability, and allelic imbalance switching in rice due to drought stress.

The indica ecotypes, IR64, an elite drought-susceptible variety adapted to irrigated ecosystem, and Apo (IR55423-01 or NSIC RC9), a moderate drought-tolerant upland genotype together with their hybrid (IR64 × Apo) were exposed to non- and water-stress conditions. By sequencing (RNA-seq) these genotypes, we were able to map genes diverging in cis and/or trans factors. Under non-stress condition, cis dominantly explains (11.2%) regulatory differences, followed by trans (8.9%). Further analysis showed that water-limiting condition largely affects trans and cis + trans factors. On the molecular level, cis and/or trans regulatory divergence explains their genotypic differences and differential drought response. Between the two parental genotypes, Apo appears to exhibit more photosynthetic efficiency even under water-limiting condition and is ascribed to trans. Statistical analyses showed that regulatory divergence is significantly influenced by environmental conditions. Likewise, the mode of parental expression inheritance which drives heterosis (HET) is significantly affected by environmental conditions indicating the malleability of heterosis to external factors. Further analysis revealed that the HET class, dominance, was significantly enriched under water-stress condition. We also identified allelic imbalance switching in which several genes prefer IR64- (or Apo-) specific allele under non-stress condition but switched to Apo- (or IR64-) specific allele when exposed to water-stress condition.

Analysis of Allelic Imbalance in Rice Hybrids Under Water Stress and Association of Asymmetrically Expressed Genes with Drought-Response QTLs.

BACKGROUND: Information on the effect of stress on the allele-specific expression (ASE) profile of rice hybrids is limited. More so, the association of allelically imbalanced genes to important traits is yet to be understood. Here we assessed allelic imbalance (AI) in the heterozygote state of rice under non- and water-stress treatments and determined association of asymmetrically expressed genes with grain yield (GY) under drought stress by in-silico co-localization analysis and selective genotyping. The genotypes IR64, Apo and their F1 hybrid (IR64 × Apo) were grown under normal and water-limiting conditions. We sequenced the total RNA transcripts for all genotypes then reconstructed the two chromosomes in the heterozygote. RESULTS: We are able to estimate the transcript abundance of and the differential expression (DE) between the two parent-specific alleles in the rice hybrids. The magnitude and direction of AI are classified into two categories: (1) symmetrical or biallelic and (2) asymmetrical. The latter can be further classified as either IR64- or Apo-favoring gene. Analysis showed that in the hybrids grown under non-stress conditions, 179 and 183 favor Apo- and IR64-specific alleles, respectively. Hence, the number of IR64- and Apo-favoring genes is relatively equal. Under water-stress conditions, 179 and 255 favor Apo- and IR64-specific alleles, respectively, indicating that the number of allelically imbalanced genes is skewed towards IR64. This is nearly 40-60 % preference for Apo and IR64 alleles, respectively, to the hybrid transcriptome. We also observed genes which exhibit allele preference switching when exposed to water-stress conditions. Results of in-silico co-localization procedure and selective genotyping of Apo/IR64 F3:5 progenies revealed significant association of several asymmetrically expressed genes with GY under drought stress conditions. CONCLUSION: Our data suggest that water stress skews AI on a genome-wide scale towards the IR64 allele, the cross-specific maternal allele. Several asymmetrically expressed genes are strongly associated with GY under drought stress which may shed hints that genes associated with important traits are allelically imbalanced. Our approach of integrating hybrid expression analysis and QTL mapping analysis may be an efficient strategy for shortlisting candidate genes for gene discovery.