Sequencing dominates latest biological research

        Having worked in biology field for 10 years, I witnessed the tremendous change in the focus researchers devote themselves to in order to solve the challenging puzzles in life sciences. Essentially, protein has given way to RNA and then RNA was surpassed by DNA. Protein dominated the stage decades ago. Western blotting, a way to detect protein, was once favored by researchers. Then RNA took the leading role. This owed to two key techniques: PCR (first developed in 1983 and awarded with Nobel prize) and microarray (nominated for Nobel prize a couple of times). Both methods use short oligos (primers in PCR and probes in microarray) to detect the amount of RNA. In this day and age, it becomes more about DNA.

        DNA has many advantages to be used to account for phenotypes. First, it is DNA that stores genetic information. Secondly, DNA encodes RNA and protein. Thirdly, DNA is more stable than RNA and protein. Protein and RNA were once heavily used to explain interesting phenotypes at a time when DNA sequencing was impractical. Nowadays, owing to the rapid progress in faster, more accurate, and more affordable DNA sequencing technology, DNA is becoming the primary choice of researchers.

       An adequate mass of tissue is favorable for DNA sequencing. However, not all cells in an organism have identical genomes. For example, tumor cells may harbor DNA mutations (the so-called tumor heterogeneity) which account for many clinical problems, such as drug resistance. Under this circumstance, single-cell sequencing would be ideal. In other occasions, single-cell sequencing is desirable because the available material is very limited. For example, in prenatal tests, only a few cells are collected from the fetus; in circulating tumor cell screening, only a few cells are shed into the bloodstream from the primary tumor; and in a forensic test, only a few cells are preserved.

        Since DNA content from a single cell is trace amount, an amplification of DNA is necessary to obtain enough DNA for sequencing. This is usually achieved by PCR amplification. However, PCR introduces extra bias, and it may not cover the whole genome. Primers (short DNA oligos used for PCR) and polymerase (the enzyme for DNA amplification in PCR) have been optimized to boost the PCR yield and the genome coverage, but the results are yet to be satisfying.  

        Recently Zong and colleagues (Science 2012: 338:6114, 1622-1626) developed a new method, MALBAC, to improve the coverage in DNA sequencing from a single cell. This method adopted multiple random primers, a DNA polymerase with strand-displacement activity, and semi-amplification process at various temperatures. By using this method, they successfully identified single-nucleotide polymorphism (SNPs) and copy-number variations (CNVs) from single human cells. This technology may prove to be a great contribution to life sciences at DNA age.