Stalled progress in reducing the malaria burden over the past few years suggests the need to develop new interventions to augment existing ones, including interventions aimed at interrupting gametocyte transmission from humans to the mosquito vector. To develop and effectively apply interventions that target gametocytes, there is a need to understand patterns of gametocytemia observed between individuals. Gametocytemia varies by host age, season, symptom status, antimalarial drugs use, and complexity of infection; however, the underlying mechanisms of this variation are not fully understood. Complexity of infection may modulate gametocytemia in P. falciparum; however, mechanisms of how clone composition influences gametocyte production are not clear. Addressing this gap requires a genotyping assay that can detect and estimate relative clone frequency of gametocytes and asexual parasites in infections. The dissertation aims were two-fold: 1) To develop an amplicon sequencing assay to genotype P. falciparum mature gametocytes; and 2) To evaluate the impact of host and parasite factors and gametocyte production in P. falciparum infections. We identified a polymorphic region of the pfs230 gene as a marker to distinguish P. falciparum mature gametocyte clones. When evaluating the impact of host and parasite factors on gametocyte production and gametocytemia, we found that more complex and high parasite density infections were more likely to produce and harbor gametocytes. The proportion of infections that produced gametocytes were similar between age-groups and between symptomatic and asymptomatic individuals, but children and asymptomatic individuals were more likely to harbor gametocytes than adults and symptomatic individuals, respectively. These findings suggest that complexity of infection and parasite density may increase gametocyte production, but additional factors such as host immunity and duration of infection may contribute to the presence or absence of gametocytes after initiation of gametocyte production. Coupled with the development of the gametocyte genotyping assay which will be an important tool for studies aimed at understanding dynamics of gametocyte production in polyclonal infections, understanding the impact of host and parasite factors on gametocyte production and gametocytemia will help explain variation in gametocytemia observed between individuals. This knowledge could inform development and effective deployment of transmission interrupting interventions.