Investigating the Role of Plant Growth Regulators for Improving Grain Size in Teff (Eragrostis Tef (Zucc.) Trotter)

Teff (Eragrostis tef) is a small grain, highly nutritious, low-risk, and warm-season annual cereal crop. Small grain size is the most important problem for teff production next to lodging. Because of this, teff experiences high post-harvest loss. Therefore, the objective of this project is to investigate the use of plant growth regulators (PGRs) to increase the grain size of teff directly and indirectly. The ethylene action inhibitor, 1-Methylcyclopropene (1-MCP) for direct effects on grain size, synthetic strigolactone (GR24) and synthetic auxin 1-
 Naphthaleneacetic acid (NAA) for indirect effects on grain size via the reduction in tillering. Teff is a semi-domesticated crop that produces an excessive number of tillers, with flowering and grain maturation occurring over an extended period. I demonstrated that application of PGRs at the appropriate time relative to panicle or plant growth stage, concentration, frequency, and interval increased grain size and thousand-grain weight (TGW). 1-MCP treatment of panicles increased grain size and TGW by 22% and 29%, and 10% and 16% with and without water stress, respectively. The effect was higher under moderate water stress. Unfortunately, to be effective, I had to treat individual panicles and treatment of whole plants was not effective. This is likely to be because of the phenology of teff flowering over an extended period (13-23 days). The difficulty of using 1-MCP at the prolonged flowering led me to look at a different strategy. Reducing tillering by GR24 and NAA treatment increase grain size and TGW by 12% and 8% and TGW by 20% and 14%, respectively. This effect is comparable with 1-MCP under normal growth conditions. Reducing tillering has also a small but positive effect on whole plant grain yield and harvest index. This is because the later tillers tend to have very small grains, with fewer tillers the plant is putting its resources into the earliest and most productive tillers/panicles. The reduction in tiller number has additional benefits like reducing lodging and seed shattering by making uniform panicle maturation. This shows targeting tillering could bring multiple trait improvements in teff. To obtain a major effect on grain size, I had to treat plants from an early stage (2-4 tillers) and for an extended period (8 times at 4-day intervals). This could make the treatment of plants in the field impractical. The positive effect of tiller reduction on grain size and TGW indicates that refocusing breeding strategies to reduce tillering or direct genetic manipulation to increase SL production would have a major positive effect on teff production. Understanding the molecular mechanism of tiller inhibition is vital to reduce tiller number via GR24 treatment, but not a lot is known in cereals, particularly in teff. To address this, I did RNA-seq upon GR24 treatment. I also investigate SL-responsive genes and other genes that are differentially expressed in actively growing tiller buds from other plants. I did not find any differential expressed genes (DEGs) 24 hours upon GR24 treatment, but I found several potential teff orthologues of plant hormone-related genes, cell cycle, sugar, transcription factor, and other genes involved in diverse functions. Cytokinin signalling and perception, auxin signalling, cell cycle, sugar, and other genes involved in diverse functions were highly expressed in growing tiller buds of teff. Particularly, HK4.2, AHP1.1, AHP1.2, IAA11, IAA30, IAA31, ARF3, ARF7.1, ARF7.2, ARF17.3, ARF21, His4, PCNA, PFPA1, Et_s981-0.21-1.path1, CTP synthase, DRM1, TPSS11, and SVP/AGL22 were expressed at high levels. The high expression of ARF7, DRM1, SVP/AGL22, and TPSS11 in growing tiller buds was reported for the first time in cereal crops. In contrast, SL transcription factor (GT1), most Arabidopsis SL responsive genes (except VLG), CK catabolism (CKX1/CKX2/CKX4/CKX9.1), transcription factor, auxin-responsive, sucrose starvation inducible (ASN1/DIN6) genes were expressed at low levels. The expression of VLG was identified for the first time in tiller buds. Exceptionally, RR4.2/RR9.2, GT1, PAP1/IAA26/IAA18, and CKX1/CKX2/CKX4/CKX9.1 genes were expressed in growing tiller buds similarly in more than three species including teff, suggesting these genes would be the primary target genes for tiller inhibition in teff. Therefore, GT1, IPA1, RR4.2/RR9.2, HK4.2, AHP1.1, AHP1.2, CKX1/CKX2/CKX4/CKX9.1, IAA11, IAA30, IAA31, ARF3, ARF7.1, ARF7.2, ARF17.3, ARF21, and PAP1/IAA26/IAA18 would be the main target genes for tiller inhibition in teff as these genes were reported functionally involved in tillering in other species. Thus, downregulating the expression of the highly expressed genes in growing tiller buds might inhibit tiller bud outgrowth. On the other hand, upregulated genes expressed at low levels in teff tiller buds might also inhibit tillering. Bud transcriptome analysis is the first study in teff and this study will be a base for teff breeders as there is a knowledge gap in a molecular study in teff particularly in tiller inhibition. Although treatment of teff in the field with 1-MCP, GR24, and NAA is unlikely to be effective to improve grain size, I have shown that research focusing on altering endogenous ethylene, SL, and auxin production or response is likely to have a positive effect on grain size and TGW with positive side benefits of reduced lodging, fewer late-flowering tillers of low productivity and a shorter season. Such plants would have major benefits both to poor farmers in Ethiopia as well as making teff more attractive to farmers outside Ethiopia seeking to diversify into specialist high-value crops