Doping has been an issue for fair sports for many years. Lance Armstrong for example won the Tour the France seven times before he was caught for the use of doping. At the end of 2012 all these seven victories were taken. The cycling world was shocked.
Athletes are constantly searching for new types of performing enhancement. Rapid advances in gene therapy enable athletes nowadays to inject themselves with performance enhancing genes, like Erythropoietin (EPO) and insulin-like growth factor 1 (IGF-1), in an almost undetectable process called gene doping. The big advantage of the use of gene doping is the easy bypassing of current detection methods. The World Anti-Doping Agency (WADA) is taking the possibility of gene doping seriously. Sooner or later, the sporting world has to deal with the phenomenon of gene doping to control athletic performance enhancement. The big problem is that no suitable detection method has been accepted for official use so far.
The TU Delft iGEM team is trying to tackle this problem by designing a suitable detection method for gene doping. In October our concept will be presented at the biggest international competition on synthetic biology, iGEM (international Genetically Engineered Machine competition). In this competition over 340 teams compete. All teams try to solve a world problem with the use of synthetic biology. This year the TU Delft is designing an efficient, secure and versatile method for the detection of gene doping.
Our novel detection method uses a dxCas9-Transposase fusion protein to target and tag the specific gene doping DNA with adapters. Only the genes with the adaptor will be sequenced with Oxford Nanopore Technologies’ next generation sequencing. With this method we create a method for targeted sequencing, which can be an efficient, secure and versatile for the detection of gene doping.
Currently, it is most likely that for the administration of gene doping, an adenovirus or a plasmid is used. Both methods bring the doping gene in the cell nucleus, where the gene will be expressed. The inserted genes however, do differ from the regular EPO genes present. Because of the size of the gene, introns must be taken out. We therefore search for the exon-exon junctions, which we do using a dxCas9 with a library of different gRNA sequences that cover all different gene doping changes. When the dxCas9 finds an exon-exon junction, the Tn5 transposase will cut the doping gene and add the sequencing adapters (Figure 1). This method for targeted sequencing will make sure that only gene doping DNA will be sequenced.
Do you want to know more about our project? Please visit our website for more information and sign up for our newsletter. We are always happy to answer questions via e-mail. You can follow us on Facebook and Twitter to stay in touch with our last sprint to the competition in October.
Get on your marks, get set and join us in the fight against gene doping.