I am studying the transcriptome of Arabidopsis. Interestingly, the 5' UTR of the latest annotation is usually too long. Here is an example (this gene is right to left). You can see both EST (orange bars) and the RNA-seq reads cover a much smaller region of the annotated 5' UTR. Unfortunately, many 5'UTR of the genes have the same problem. Is there a way to fix that? I was wondering if any assembly tools can help me to get the correct 5'UTR ranges. Thanks!!!
- Genes, particularly transcribed genes in Eukaryotes, can have alternate start and stop sites (and splicing). Alternate transcriptional start sites may be found in different tissues or at different developmental periods due to alternate transcription factor binding sites being used. The only reliable way to determine the transcriptional start and stop sites is by read coverage.
- Genome annotations are generally the product of bioinformatic pipelines and are mostly predictions based upon computational models. Different gene-finders may call different boundaries; for example, in eukaryotic genomes, an exon may be assigned to the wrong up/down-stream gene. Arabadopsis, as model organism, has had it's gene boundaries considerably refined/corrected/validated, but even so, there could still exist errors in the annotation or, more likely, the boundaries indicated are the most common start/stop sites (see point 1 above).
In short, your UTRs are whatever your experimental data (reads) show it to be. Don't worry too much about it not matching up to the standard annotation boundaries.