Universal endonuclease – cutting edge technology
Abstract
Gene technology is driven by the use of restriction endonucleases. Yet, constraints of limited sequence length and variation recognized by available restriction enzymes pose a major roadblock for synthetic biology. We developed the basis for universal restriction enzymes, primarily for routine cloning but also with potential for in-vivo applications. We use a nucleotide cleavage domain fused to a binding domain, which recognizes a programmable adapter that mediates DNA binding and thus cleavage. As adapter we use readily available modified oligonucleotides, as binding domain anticalins, and as cleavage domain FokI moieties engineered for heterodimerization and activity. For application, this universal enzyme has merely to be mixed with the sequence-specific oligonucleotide and the target DNA. Binding and release are addressed by thermocycling. We provide concepts for in-vivo applications by external adapter delivery and activity regulation by photo switching. Additionally, an argonaute protein is engineered towards a DNA endonuclease.
Wiki
https://2009.igem.org/Team:Freiburg_bioware
Achievements
- Finalist
- Gold Medal
- Best Poster
- Best New BioBrick Part or Device, Engineered
Meet The Team
Primary PI
Instructors
- Sven Hagen
- Tobias Baumann
- Dr. Katja Arndt
- Janina Speck
Students Members
- Anika Lückoff
- carolin Nieder
- Christoph Deeg
- Dieter Huber
- Hannes Breddermann
- Isabel Lohrmann
- Julia Michely
- Laura Eichstädter
- Manuel Hiß
- Maximilian Müller
- Paul Staab
- Rüdiger Trojok
- Sarah-Katharina Bambach
- Sascha Duttke
- Timo Lorenz