Fusion transcripts are the result of chromosomal abberations, where former distant genes are then fused together, sharing one transcriptional start site and creating a novel protein with new, unwanted characteristics. This fusion genes are acting as key elements (enhancers) in certain cancer types. Hence the detection of chimeric genes is often investigated in cancer biology. On RNA-Seq level this is usually done by investigating reads, where their two ends are detected to align to chromosomal locations with a large distance. Also paired-end reads are taking into account. Here the two reads of one pair can align to a different position in the genome resulting in large distances between the locations of the the two reads.
One obstacle in calling chimeric transcripts is the abundance of many false-positive fusion events in the raw fusion gene list. This can be due to several reasons. Prominent examples for events that give false positive fusion transcripts are pseudogenes, paralogs, naturally overlapping genes or simply “real” fusions, which had been found in samples from healthy donors, with no pathological outcome. We are using tools, which take those critical features into account to filter the preliminary list of fusion events to a final list of candidate fusion transcripts/genes. While this approach ellimates a high percentage of the initial candidate list, it is still a very sensitive approach for fusion detection.
We then carefully investigate all fusion events by visual examination of all raw reads mapping to the breakpoint regions of the fusion (please see example on the right).
No result counts, if not presented in the best way. We are aiming for high-quality figures. We provide high-resolution images and additionally pdf versions of your graphics, which enable you to manipulate colors, text and many other options. Please see an example video here.
In case you want to contract it’s biology to analyse your Fusion detection project, we will divide the whole process into 3 steps, with you choosing, which level of analysis you need:
While physically there , fusion transcripts can be transcribed at very low level in the samples you are investigating. Therefore it is alsways recommended to archieve a sufficient read depth when evaluating chimeric transcripts. Talk with us upfront.
Data preparation and alignment
Heat our workstations: Data quality assessment, read manipulation (trimming, filtering) and alignment of your RNA-Seq reads. Please have a look at the RNA-Seq section in our Portfolio for more information on this steps.
Fusion events: detection, annotation, filtering and visualization
All you need to detect and interpret chimeric transcripts in your samples.
We are analyzing RNA-Seq data for fusion transcript detection from all mayor next-generation sequencers from Illumina or Ion-Torrent. We can start from files in FASTA, FASTQ, unaligned BAM files or SRA format.
For getting an overview of the detected fusion transcripts there is nothing better than a Circos Plot. We are adapting this plot type to your needs, focusing on specific regions in the genome if needed. Additionally we can add more tracks in the plot, for example derived from the actual expression data or from various databases.
We also supply you with genomic plots of the exact breakpoint of your fusion events. This both focused on the breakpoint, showing the overlapping single end reads and also more zoomed out, for visualization of paired end reads supporting your fusion events.