Engineering RNA logic with synthetic splicing ribozymes

Reusable components, such as logic gates and code libraries, simplify the design and implementation of electronic circuits and computer programs. The engineering of biological systems would benefit also from reusable components. In this thesis, I show the utility of splicing ribozymes for the biological engineer. Ribozymes allow the engineer to manipulate existing biological systems and to program self-modifying RNA systems. In addition, splicing ribozymes are easy to engineer, malleable, modular, and scalable. I used the model ribozyme from Tetrahymena to explore the principles behind engineering biological splicing systems in vivo. I show that the core ribozyme is modular and functions properly in many different contexts. Simple base pairing rules and computational RNA folding can predict splicing efficiency in bacterial cells. To test our understanding of the ribozyme, I generated synthetic ribozymes by manipulating the primary sequence while maintaining the secondary structure. Results indicate that our biochemical understanding of the ribozyme is accurate enough to support engineering. Splicing ribozymes can form core components in an all-RNA logic system. I developed biological transzystors, switches analogous to electrical transistors. Transzystors can use any trans-RNA as input and any RNA as output, allowing the genetic reading of RNA levels. I also show the ribozyme can write RNA using the trans-splicing reaction.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Electrical Engineering and Computer Science.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2008

Author dc:creator

• Che, Austin, 1979-

Advisor dc:contributor.advisor

• Thomas F. Knight, Jr.

• Electrical Engineering and Computer Science.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/47786

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/47786