This review highlights the developments, accomplishments and challenges of integrated μPADs, including sample pretreatment, signal transduction/amplification and results output.
High-quality whole-genome
amplification (WGA) of individual cells
is the primary step for characterizing the genetic information on
single cells in biology and medicine. As the most popular single-cell
WGA method, multiple displacement amplification (MDA) is often plagued
by the nonuniform amplification. The droplet MDA has been an innovative
tool to solve this dilemma by mitigating the amplification bias and
increasing the genomic coverage. Despite these advantages, the time-consuming
droplet generation process, the waste of small volume samples and
the difficulty of parallel operation for multiple single-cell samples
remain major obstacles. Herein, we introduce a centrifugal-driven
droplet generation method for rapid and convenient generation of uniform
droplets from a relatively small volume sample (5 μL) in 60s
with more than 98% sample utilization. We have performed quantitative
digital droplet PCR using this method, demonstrating its capability
of amplifying nucleic acids at the single-molecule level. Single-cell
centrifugal-driven droplet MDA (cd-MDA) has also been conducted for
single-cell sequencing, achieving uniform amplification and broad
genomic coverage. With the single-molecule sensitivity, minimum sample
waste, high genomic coverage, and excellent sequencing evenness, this
centrifugal-driven droplet generation method is promising for convenient
and scalable use in digital PCR and single-cell whole-genome research.
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