Measuring and Detecting Transcription
Transcription can be measured and detected in a variety of ways:
- Nuclear Run-on assay: measures the relative abundance of newly formed transcripts
- RNase protection assay and ChIP-Chip of RNAP: detect active transcription sites
- RT-PCR: measures the absolute abundance of total or nuclear RNA levels, which may however differ from transcription rates
- DNA microarrays: measures the relative abundance of the global total or nuclear RNA levels; however, these may differ from transcription rates
- In situ hybridization: detects the presence of a transcript
- MS2 tagging: by incorporating RNA stem loops, such as MS2, into a gene, these become incorporated into newly synthesized RNA. The stem loops can then be detected using a fusion of GFP and the MS2 coat protein, which has a high affinity, sequence-specific interaction with the MS2 stem loops. The recruitment of GFP to the site of transcription is visualised as a single fluorescent spot. This remarkable new approach has revealed that transcription occurs in discontinuous bursts, or pulses (see Transcriptional bursting). With the notable exception of in situ techniques, most other methods provide cell population averages, and are not capable of detecting this fundamental property of genes.
- Northern blot: the traditional method, and until the advent of RNA-Seq, the most quantitative
- RNA-Seq: applies next-generation sequencing techniques to sequence whole transcriptomes, which allows the measurement of relative abundance of RNA, as well as the detection of additional variations such as fusion genes, post-translational edits and novel splice sites
Read more about this topic: Transcription (genetics)
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