Is it possible to simulate DNA

Health Industry BW

Enormous advances have been made in recent years in the possibilities of deciphering the genome: DNA sequencing techniques have been developed under high pressure according to the motto "faster, further, more cost-effective". With today's methods, it is already possible to cope with larger projects in a short time. However, the error rate is still very high, which is why a sample usually has to be sequenced several times in order to be able to determine the correct sequence. The junior professor Dr. Maria Fyta and her research team at the Institute for Computational Physics at the University of Stuttgart and colleagues in Sweden and Brazil have now found out in simulation calculations that special chemical modifications in the nanopores of sequencing devices would make the process much more error-free and therefore more efficient.

The decoding of the human genome, completed in 2001 and involving many scientists around the world, took around 13 years and cost more than $ 3 billion. Nowadays, such a project would lead to results much faster and cheaper in just a few days. In particular, the Next Generation Sequencing (NGS) methods have enormously expanded the possibilities of analyzing genes and genomes, even for smaller institutes and biotech companies. However, a high error rate must often be expected with the high-tech processes. Although this can be compensated for by reading the sequences multiple times, it understandably reduces the efficiency. In addition, enormous amounts of data are produced, which have to be evaluated in a time-consuming and computational manner.

Research is therefore being carried out in many places into ways of reducing the errors in the decryption of nucleic acids and, at the same time, increasing the accuracy. The junior professor Dr. Maria Fyta and her working group from the Institute for Computational Physics at the University of Stuttgart have been working on a sub-project of the Collaborative Research Center 716 (“Dynamic simulation of systems with large particle numbers”) for some time together with their colleagues Prof. Dr. Ralph Schleicher in Sweden and Prof. Dr. Rodrigo Amorim in Brazil with methods that make the sequencing process even more efficient. In their simulation calculations, the scientists look at the analysis of genetic information in nanopores. “Experiments with nanopores with regard to sequencing methods have been going on for 20 years,” explains Fyta. “Very small holes in the nanometer range are drilled in various materials. Biological nanopores, for example transmembrane proteins, and solids such as silicon nitride or graphene are used for this purpose. Electrophoresis then takes place in the nanopores. "

In nanopores, nucleotides have different electronic properties

The analysis of genetic information with the help of nanopores is counted as part of the so-called third generation sequencing. This includes the most modern high-tech processes in which the base sequence is determined by detecting individual molecules. With nanopore sequencing, it should theoretically be possible to decode a complete human genome in one day for an acceptable price. Devices based on nanopores have also been available for some time. “For example, Oxford Nanopore Technologies offers sequencers that are almost as small as a USB stick and can read several thousand base pairs,” reports Fyta. "However, the process is still very flawed and can therefore currently not be used in medicine."

In nanopore sequencing, as in other sequencing techniques, nucleic acid fragments of different sizes are separated electrophoretically in order to be able to determine the sequence of the base components. For this purpose, the DNA to be examined is placed in saline solution and an electrical voltage is then applied to the nanopores. Gold electrodes are currently used in almost all techniques based on nanopores. An electric field is formed in the vicinity of the pores, in which the negatively charged DNA molecules move and are drawn very quickly through the pore. The sequence of the nucleotides can be determined by detecting differences between the four different bases: Although they are chemically very similar, they differ in their electronic properties. "With the method we examined, you can read the DNA code by measuring the so-called cross current through the gold electrodes," explains the junior professor. "And it is actually the case that you get different electrical signals, depending on which nucleotide passes through the pore."

Coating made from modified diamondoids

The Stuttgart scientists have been carrying out such experiments for some time now, using simulation calculations to look primarily at the behavior of nucleic acids in the cross-flow. "At some point we asked ourselves whether we could perhaps better differentiate between the individual bases if the gold electrodes in the pores were coated with diamond-like molecules," says Fyta. And indeed: the scientists were able to observe that the differences between the nucleotides in the cross-flow became even more specific. “They were easier to distinguish because the nucleic acid in the coated nanopore forms hydrogen bonds between nucleotides and the diamond - each of the four nucleotides in a slightly different way,” reports the computer physicist. "The specificity of the hydrogen bridge formation in the cross-flow can be seen relatively clearly with the appropriate methods, and the four nucleotides can be distinguished from one another much better in our calculations than without a diamond coating." 1

Diamondoids can be produced and specifically modified by chemists, as Fyta says. “Tiny molecules from diamond-like cages are used for the coating,” explains the scientist. “These diamond molecules are surrounded by a termination made up of hydrogen molecules and other functional groups of atoms that are important for the formation of hydrogen bonds with the nucleotides.” So the technology itself should work. However, what was still missing in the calculations of the Stuttgart scientists are the external test conditions - the salt and the water, as Fyta says: “We still have to simulate what the salt solution will contribute to this. And so far we have assumed individual nucleotides in our calculations. But of course we also have to consider what happens when you use more nucleic acids. "

Simulation calculations now also for the system as a whole

For the simulations, the computer physicists did not choose a model that looks at the system in a classic way, but rather a quantum mechanical approach in which the complete system of DNA, nanopore and electrodes is included. “We decided on this approach because the interactions can be estimated more precisely,” explains the junior professor. Such a simulation takes several days even on high-performance computers. But in Baden-Württemberg there are many supercomputers on which university institutes have computing time, says Fyta.

In the next few months, the scientists want to look at the experimental conditions that have not been taken into account so far, as well as the system as a whole: “Because a single nucleobase in a single nanopore does not naturally occur in any laboratory experiment. That is why we want to test long nucleic acids in larger systems in the near future, ”says the scientist. At the very end of the studies - if all tests continue to be successful - the development of devices with diamond-coated gold electrodes in the nanopores should of course take place. Before that, of course, the practical tests in the laboratory would have to be done.

1 Sivaraman, Ganesh; Amorim, Rodrigo G .; Scheicher, Ralph H .; Fyta, Maria: Diamondoid-functionalized gold nanogaps as sensors for natural, mutated, and epigenetically modified DNA nucleotides. In: Nanoscale, 2016,8, 10105-10112, DOI: 10.1039 / C6NR00500D