English: How PTA works. PTA may be performed directly from single cells (collected by FACS, microfluidic or other methods), multiple cells, or ultra-low inputs of DNA (>4 pg– 10 ng). After denaturation, random primers (6- to 9-mers) are annealed. Extension with phi29 and a proprietary nucleotide pool results in amplicons of ~250 to >1,500 bp in length. The relatively small size of these amplicons favors subsequent priming off the primary template, thereby limiting the exponential propagation of biases and errors in daughter molecules. In addition, the PTA chemistry suppresses the formation of experimental artifacts such as chimeric molecules and non-specific priming.2 PTA reaction products are double-stranded, and may be converted to libraries for multiplexed sequencing on Illumina® or other short-read platforms without fragmentation.
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