Einzelmolekülsequenzierung

Einzelmolekülsequenzierung is a powerful technology that allows for the determination of the nucleotide sequence of single DNA or RNA molecules. Unlike traditional sequencing methods that rely on amplifying DNA fragments before sequencing, single-molecule sequencing directly reads the sequence of individual molecules. This bypasses the biases introduced by amplification steps, potentially leading to more accurate and comprehensive results.

The technology has evolved significantly over the years, with different approaches being developed. Early methods often involved single-molecule fluorescence in situ hybridization (smFISH) or primer extension assays. More recently, advancements in nanopore sequencing and single-molecule real-time (SMRT) sequencing have become prominent. Nanopore sequencing, for example, involves passing a single DNA or RNA molecule through a protein or solid-state nanopore. As the molecule translocates, changes in electrical current are detected, which correspond to specific nucleotides. SMRT sequencing, on the other hand, uses fluorescently labeled nucleotides and observes their incorporation by DNA polymerase in real-time.

The applications of Einzelmolekülsequenzierung are broad and expanding. It is crucial for de novo genome assembly, especially for complex and repetitive regions that are challenging for short-read sequencing. It also enables the detection of structural variations, epigenetic modifications (like methylation), and direct RNA sequencing without the need for conversion to cDNA. Furthermore, its ability to sequence long molecules can improve the resolution of transcriptomics, providing insights into full-length transcripts and alternative splicing events. The field continues to advance, promising even greater resolution and accessibility in the future.