1 research outputs found
Precise Expression Profiling by Stuffer-Free Multiplex Ligation-Dependent Probe Amplification
In
systems biological studies, precise expression profiling of
functionally important gene sets is crucial. Real-time polymerase
chain reaction is generally used for this purpose. Despite its widespread
acceptance, however, this method is not suitable for multiplex analysis,
resulting in an inefficient assay process. One alternative technology
in the spotlight is multiplex ligation-dependent probe amplification
(MLPA). But MLPA depends on length-based discrimination of amplified
products, which complicates probe design and compromises analysis
results. Here, we devised a variation of MLPA that utilizes conformation-sensitive
capillary electrophoresis, and demonstrated the simplicity of the
probe-design process and improved precision of the assay in analyses
of 33 <i>Escherichia coli</i> metabolic genes and 16 <i>Caenorhabditis elegans</i> longevity-related genes. The results
showed that relative expression could be quantitatively measured over
a relevant dynamic range by using similar-sized probes. Importantly,
the improved precision compared to conventional MLPA promises a wider
application of this method for various biological systems