Doctor of Philosophy, Case Western Reserve University, 2023, Biology

Tylosema esculentum (marama bean) is an underutilized legume, long considered as a local potential crop due to its rich nutritional value. The reference plastome and mitogenome were assembled using a hybrid method with both Illumina and PacBio data. The diversity was explored with the WGS data of 84 samples from various geographic locations in Namibia and Pretoria. Phylogenetic analysis revealed two cytotypes with distinct plastomes and mitogenomes with differing levels of variability. Deep sequencing has identified heteroplasmy with both types of organellar genomes present, albeit one at a very low frequency. The inheritance of this complex of organellar genomes appears to be fairly constant, providing a conundrum of how the two genomes co-exist and are propagated through generations. The type 1 mitogenome has two autonomous rings with a total length of 399,572 bp, which can be restructured into five smaller circular molecules through recombination on 3 pairs of long direct repeats. The type 2 mitogenome contains a unique 2,108 bp sequence, which connects distant segments to form a new structure consisting of three circular molecules and one linear chromosome. This increased the copy number of nad9, rrns, rrn5, trnC, and trnfM. The two mitogenomes differed at another 230 loci, with only one nonsynonymous substitution in matR. cpDNA insertions were concentrated in one subgenomic ring of the mitogenome, including a 9,798 bp long fragment that contains potential psbC, rps14, psaA, and psaB pseudogenes. The two types of plastomes range in length from 161,537 bp to 161,580 bp, differing at 122 loci and at a 230 bp inversion. The chloroplast genes rpoC2, rpoB, and ndhD were found to be more diverse than other genes in marama plastome. 21.6 Gb PacBio HiFi data was assembled using Canu v2.2 into an unphased assembly of 1.24 Gb. k-mer analysis indicated that marama may be ancient tetraploid with an estimated genome size of only 277 Mb. The generated assembly has an N50 v (open full item for complete abstract)

Committee: Christopher Cullis (Advisor); Hillel Chiel (Committee Chair); Peter Zimmerman (Committee Member); Jean Burns (Committee Member); Sarah Bagby (Committee Member) Subjects: Bioinformatics; Genetics; Plant Biology

Master of Sciences, Case Western Reserve University, 2011, Biology

The intergenic spacer (IGS) of the large rDNA gene array has been characterized in organisms across the domain Eukarya. A common characteristic of this region is length heterogeneity, not only between conspecific individuals, but within individual organisms. In most species, this heterogeneity has been found to result from one or more clusters of subrepeats, each containing a varying number of subrepeat units. Length heterogeneity has previously been observed in the IGS of the undomesticated legume Tylosema esculentum (commonly called marama). This locus was sequenced and the region containing subrepeat clusters compared amongst 21 individuals. The subrepeat clusters contributing to IGS length heterogeneity are identified, and the organization of these clusters within the marama IGS is compared to that of other species. The findings presented here suggest that the subrepeat region in marama is smaller and more complex than in the analogous regions of other species.

Committee: Christopher Cullis Ph.D. (Advisor); Emmitt Jolly Ph.D. (Committee Member); Barbara Kuemerle Ph.D. (Committee Member) Subjects: Agriculture; Bioinformatics; Biology; Botany; Genetics; Molecular Biology; Plant Biology; Plant Sciences

Master of Sciences, Case Western Reserve University, 2011, Biology

The 5S ribosomal DNA regions of genomes have recently come to attention for genetic studies due to its highly variable spacer region in conjunction with a conserved gene region. It is especially of use for Tylosema Esculentum (Marama) due to the lack of sequencing data available for the species. This study focused on the variation of the 5S region of marama as compared to other species and sought to find explanations for this deviance. Specific marama 5S primers were generated to generate simpler bands to analyze. The resulting sequences showed large variation within each class size, not seen in other species as well as considerable variation between spacer regions.

Committee: Christopher Cullis Ph.D. (Advisor); Claudia Mizutani Ph.D. (Committee Chair); Emmitt Jolly Ph.D. (Committee Member); Barbara Kuemerle Ph.D. (Committee Member) Subjects: Biology; Botany; Genetics