DNA sequencing

In DNA sequencing, biochemical methods are used to determine the sequence of nucleotides (DNA base molecule with sugar and phosphate) in a DNA molecule. The most widely used method is the Sanger chain termination method. Since DNA is composed of four different bases, four different approaches are made.

Each approach contains the DNA to be sequenced, a primer (starting molecule for sequencing), DNA polymerase (enzyme that extends DNA) and a mixture of all four required nucleotides. However, in each of these four approaches a different base is chemically modified in such a way that it can be incorporated but does not provide a point of action for the DNA polymerase. This then leads to chain termination. This method produces DNA fragments of varying lengths, which are then chemically separated by so-called gel electrophoresis according to their length. The resulting sorting can be translated into the sequence of nucleotides in the sequenced DNA section by labeling each base with a different fluorescent color.

DNA Hybridization

DNA hybridization is a molecular genetic method used to prove the similarity between two single strands of DNA of different origin. This method makes use of the fact that a DNA double strand is always composed of two complementary single strands. The more similar both single strands are to each other, the more bases form a solid connection (hydrogen bonds) with the opposite base or the more base pairs are formed.

No base pairings will occur between sections on the two DNA strands that have a different base sequence. The relative number of compounds can now be determined by determining the melting point at which the newly formed DNA double strand is separated. The higher the melting point, the more complementary bases have formed hydrogen bonds to each other and the more similar the two single strands are. This method can also be used to detect a specific base sequence in a DNA mixture.For this purpose, artificially formed DNA fragments can be labeled with (fluorescent) dye. These then serve to mark the corresponding base sequence and can thus make it visible.