This research explored the role of the OsWOX9A protein in zygotic and somatic embryogenesis and improvements to synthetic apomixis in rice. To enhance synthetic apomixis and investigate its role in early embryogenesis, OsWOX9A was studied using the Arabidopsis egg cell promoter, pDD45, in rice. Despite initial expectations, pDD45::OsWOX9A alone failed to initiate parthenogenesis. Next, the rice egg cell promoter, pOsECA1.1, was evaluated to try and improve the efficiency of synthetic apomixis in rice. pOsECA1.1 was shown to have specific egg cell expression both in rice and Arabidopsis and provided a substantial increase in the efficiency of synthetic apomixis in rice (Vernet et al., 2022). To further study OsWOX9A, the glucocorticoid receptor (GR) dexamethasone-inducible system was employed. OsWOX9A-GR induction in rice, revealed distinct root phenotypes, including corkscrew-like growth patterns, inhibition of lateral roots and enhanced tissue greening, all of which suggest OsWOX9A plays a role in cytokinin signaling and or regulation. Based on this result and prior data that OsBBM1-GR can induce callus but fails to enable shoot regeneration, the two transcription factors were combined to assess whether complete somatic embryogenesis could be achieved (Khanday et al., 2023). Simultaneous induction of both transcription factors did not achieve this but rather induced strong proliferation of embryogenic calli. Complete and highly efficient somatic embryogenesis could likely be achieved with modifications and fine tuning of this system. OsWOX9A-GR was also used to elucidate the genetic pathways in rice early embryogenesis. Using RNA-seq, several direct transcriptional targets of OsWOX9A were confirmed including OsWOX2, OsPIN5b, and OsKitaake12g221200. This presents further candidates for study in zygotic embryogenesis, as well as other avenues for improvement of somatic embryogenesis and synthetic apomixis biotechnologies.

Treated sewage, stormwater runoff and industrial waste are sources of chemical contamination in coastal sediments. Although studies of sedimentary chemistry have resulted in stringent laws on discharging toxic chemicals into the sea, the more toxic chemicals can have lasting effects on the environment.

Polychlorinated biphenyls (PCBs) are an important class of industrial pollutants that areubiquitous in the marine and freshwater environments (NAS, 1979). Although the “planar” or“non-ortho” PCB congeners are present in much lower concentrations than other PCBcongeners, they can be responsible for much of the dioxin-like (TCDD) toxicity in the marineecosystem. Further, their environmental fate may be different from other PCBs. Theirdetermination is, therefore, crucial for assessment of contaminant status of environmentalcompartments. Mono-ortho PCBs and polychlorinated naphthalenes (PCN) although less potent,also exhibit TCDD-like toxicity. Yet, only total PCBs are usually reported and the levels of non-and mono-ortho PCBs relative to total PCBs have rarely been studied and data on PCN areuncommon in oceanic samples (Jarnberg et al., 1993). Among the known sources, municipalwaste dischargers could be a significant contributor of these synthetic compounds to the ocean(i.e., Venkatesan, 1998; Zeng and Venkatesan, 1999).

AbstractGenome-wide Redistribution of siRNAs in Rice Gametes and Zygotes

Gametes and zygote constitute a critical stage in the life cycle for all sexual organisms, including plants. For angiosperms, the seed is the basic reproductive unit, which is produced by double fertilization. The male gametophyte, pollen, contains two sperm cells. One of the sperm cells fuses with the egg cell to produce the zygote, which will develop into the embryo and the next sporophytic generation. The other sperm cell fuses with the diploid central cell to produce the endosperm, a nutritive tissue that supports the growth of the embryo or germinating seedling. Seeds — either the endosperm (e.g., rice, maize, wheat, and other cereals) or the embryo (e.g., soybean, peanut, and other pulses) — directly or indirectly account for most of the calorie human consume, and seeds are the product of double fertilization. Thus, understanding the biology of gametes and zygote has broad applications in biotechnology and agriculture. In this dissertation I detail an aspect of the biology of rice gametes and zygote: the small RNA transcriptome and its implications regarding epigenome and plant development. In the first chapter I introduce 1) zygotic genome activation in plant, a process during which the fertilized egg cell transition from the gametic cell fate to the embryonic and totipotent cell fate; and 2) epigenomic reprogramming that was predicted or reported before and after fertilization in angiosperms. In the second chapter, I present detail protocols for isolating rice gametes, which were used to generate data presented in later chapters of this dissertation. In the third chapter, I present results of a study characterizing the small RNA transcriptomes and DNA methylomes of rice gametes. The results indicate that a genome-wide redistribution of 24-nt siRNAs has occurred in rice gametes, which was unexpected and not predictable from our canonical understanding of siRNA functions. However, in both gametes, the patterns of CHH methylation, a strong indicator of RNA-directed DNA methylation, remain similar to each other and to vegetative tissues. These findings are suggestive to unexplored roles of gamete small RNAs. In the fourth chapter, I present results of a study characterizing the small RNA transcriptome of rice zygotes. The results indicate that wide-spread redistribution of siRNAs occurred in zygote, and newly detected siRNA loci in zygote have a distribution similar to that of canonical siRNA loci detected in embryo and seedling. In addition, zygote siRNA loci, but not egg siRNA loci, were associated with hypermethylation in mature embryo. These findings suggest that resetting of the gametic epigenome towards the canonical vegetative pattern is initiated in the zygote, setting the stage for RdDM activity during embryogenesis before the first embryonic division. Lastly, I briefly summarize key findings of my dissertation and discuss possible future experiments in the final chapter of this dissertation.

Plant-associated microbes form complex interactions within their microbial community as well as with the host, affecting processes such as nutrient and carbon cycling, abiotic stress protection, and disease resistance. Understanding the interactions of the host and host-associated microbiomes could potentially aid plant health in a variety of ways and is therefore of interest to the plant science community. Rice is a unique system to study plant-microbe interactions because of flooded conditions in which it is cultivated. This anoxic environment leads to a host of anaerobic microbial taxa associating with the rice host which harbor functions dependent on the anoxic conditions, including methanogenesis. In this thesis, I describe three studies where we profile the taxonomies and functions of rice-associated microbiomes and demonstrate how the knowledge gained from these methods can be applied in discovering microbial isolates that are beneficial to the plant host.In the first chapter, I present a study that characterizes the microbiomes of a high- and a low-methane-emitting rice cultivar throughout the growing season. We found that the high methane emitting cultivar had a higher relative abundance of methanogenic archaea in the rhizosphere, and additionally had a greater abundance of taxa associated with fermentation, which could be responsible for producing methanogenic precursor molecules. In the second chapter, I present a study that characterizes the effects of drought and recovery on the microbiome. We found that a Streptomyces taxa became the most abundant member of the endosphere community during and after drought, and that this member was prevalent in rice samples from various locations. We cultured a corresponding isolate and demonstrated that it had similar qualities to the Streptomyces from the study (it was endophytic and increased in relative abundance under drought stress) and promoted root growth, demonstrating the enrichment of this microbe under drought could potentially mitigate drought effects. In the third chapter, I present a study demonstrating the effects of nitrogen on the microbiome in three different soils by profiling both the taxonomies and functions through genome-resolved metagenomics. We found that the response of rice-associated microbiomes to the nitrogen fertilizer was largely soil specific, though there were general increases in Gammaproteobacteria and decreases in Actinobacteria in response to nitrogen. Profiling the metagenome demonstrated that nitrogen fertilizer increased the abundance of genes related to aromatic compound degradation, fermentation, and methanogenesis. We also recovered 60 metagenome assembled genomes, which to our knowledge is the first such dataset of its kind in rice, and can be a valuable resource to further understanding the dynamics of microbes associating with rice.

Graph-structured data appears in many modern applications like social networks, sensor networks, transportation networks and computer graphics. These applications are defined by an underlying graph (e.g. a social graph) with associated nodal attributes (e.g. number of ad-clicks by an individual). A simple model for such data is that of a graph signal--a function mapping every node to a scalar real value. Our aim is to develop signal processing tools for analysis of such signals de- fined over irregular graph-structured domains, analogous to classical Fourier and Wavelet analysis defined for regular structures like discrete-time sequences and two-dimensional grids.

In this work, we start by reviewing the notion of a Graph Fourier Transform (GFT), which has been defined in the literature for graph signals. We examine the spatial and spectral features of circulant graphs, which accommodate linear shift-invariant operations. We describe fundamental operations such as shifting, sampling, graph-reconnection and linear filtering for signals on circulant graphs and derive associated sampling and graph-reconnection theorems. We also develop wavelet filter bank structures for multi resolution analysis of large-scale graphs.

We present a method to decompose an arbitrary graph into a linear combination of circulant graphs. This helps extend fundamental operations such as sampling, filtering and multi resolution filter banks to general graphs. We present an application in the area of graph semi-supervised learning where some of the existing algorithms can be viewed as suitably designed filters defined in the GFT domain. We propose a wavelet regularized learning algorithm and evaluate the performance on some real-world datasets.