Document Type

Thesis - Open Access

Award Date


Degree Name

Master of Science (MS)


Biology and Microbiology

First Advisor

Ruanbao Zhou


biotechnology, metabolic engineering, microbiology, synthetic biology


Growing energy demand and rising levels of greenhouse gases has put massive strain on the global environment. Alternatives to fossil fuels are being developed in an attempt to curb climate change. Biotechnology has made large strides in order to create a completely renewable energy source by genetically modifying microbes to produce biofuels and other “green” high value compounds. In this thesis project, (1) E. coli ATCC9637 (E. coli W) was genetically modified to produce bioethanol from beet juice which contains mainly sucrose. The ethanol productivity by engineered E. coli W was 18.8 mg/L/H/OD600.
(2) Cyanobacterium Anabaena sp. PCC7120 was successfully engineered to produce and secrete biofuel ethanol using CO2, water and sunlight.
(3) Another attempt in this study was made to increase production of bioethanol in ethanol-producing Anabaena strain by introducing a supplementary CO2-fixing photorespiratory bypass pathway. Although Ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) is responsible for the majority of carbon assimilation on Earth, RubisCO has poor specificity between CO2 and O2 which can lead to photorespiration and ultimately the loss of fixed carbon and nitrogen. The 3-hydroxypropionate (3-HPA) bypass was introduced into ethanol-producing Anabaena strain in an attempt to increase CO2-fixation and then increase ethanol production. Introduction of the 3-HPA bypass into Anabaena showed only a marginal increase photosynthetic activity and a decrease in growth rate. Additional genetic manipulation would be required to alleviate bottlenecks and toxic intermediates in order to create a fully functioning supplementary CO2-fixation pathway in Anabaena 7120.

Library of Congress Subject Headings

Ethanol as fuel.

Synthetic biology.

Biomass energy.


Sugar beet.



Includes bibliographical references (pages 95-102)



Number of Pages



South Dakota State University


Copyright © 2016 Nathanael Braselton