Single cell sequencing

Single cell sequencing examines the sequence information from individual cells with optimized next generation sequencing (NGS) technologies, providing a higher resolution of cellular differences and a better understanding of the function of an individual cell in the context of its microenvironment.

A typical human cell consists of about 6 billion base pairs of DNA and 600 million bases of mRNA. With such huge amount of sequence, it is expensive and time-consuming to sequence by traditional Sanger sequencing. By using deep sequencing of DNA and RNA from single cell, cellular functions can be investigated extensively. Like typical NGS experiments, the protocols of a single cell sequencing generally contain the following steps: isolation of single cell, nucleic acids extraction and amplification, sequencing library preparation, sequencing and bioinformatic data analysis. It is more challenging to perform single cell sequencing in comparison with sequencing from cells in bulk. The minimal amount of starting materials from a single cell make degradation, sample loss and contamination exert pronounced effects on quality of sequencing data. In addition, due to the picogram level of the amount of nucleic acids used, heavy amplification is often needed during sample preparation of single cell sequencing, resulting in the uneven coverage, noise and inaccurate quantification of sequencing data.

Recent technical improvements make single cell sequencing a promising tool for approaching a set of seemly inaccessible problems. For example, heterogeneous samples, rare cell types, cell lineage relationships, mosaicism of somatic tissues, analyses of microbes that cannot be cultured, and disease evolution can all be elucidated through single cell sequencing. Single cell sequencing was selected as the method of the year 2013 by Nature Publishing Group.

Single cell DNA genome sequencing involves isolating a single cell, performing whole-genome-amplification (WGA), constructing sequencing libraries and then sequencing the DNA using a next-generation sequencer (ex. Ion Torrent, Illumina). It can be used in metagenomics studies and when sequencing the first time from novel species. In addition, it can be united with high throughput cell sorting of microorganisms and cancer. One popular method used for single cell genome sequencing is multiple displacement amplification and this enables research into various areas such as microbial genetics, ecology and infectious diseases. Furthermore, data obtained from microorganisms might establish processes for culturing in the future. Some of the tools that can be used for single cell genome sequencing include: SPAdes, IDBA-UD, Cortex and HyDA.

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