Methods for iPS Cell Reprogramming

Recent studies have demonstrated the ability to reprogram adult fibroblasts into pluripotent stem cells by introducing key factors using viral vector technologies. Induced pluripotent stem cells (iPS cells) hold much promise for the advancement of stem cell research.

This breakthrough research was performed in the laboratory of Dr. Shinya Yamanaka at Kyoto University using retroviruses to introduce key stem cell factors: Oct3/4, Klf4, Sox2, and c-Myc. Other studies have shown that two other factors may also be useful in iPS generation: NANOG and Lin-28. More recent studies have shown that these genes can be introduced by other methods including adenoviruses or plasmids, with the idea that non-viral methods may be safer than viral methods. However, retrovirus remains the method of choice because of high efficiency levels.

More recently, researchers have developed a single lentiviral vector containing all four original stem cell factors (lentivirus is a subclass of retrovirus). The pLentG-KOSM vector generates iPS cells with at least 10-fold higher efficiency than the original methods, in part because it is easier to infect cells with a single virus than with a pool of multiple viruses. If even one virus in a pool does not infect the target cell, the efficiency of reprogramming can drop significantly. Lentiviral vectors have also proven to be effective in transducing brain, liver, muscle, and retina in vivo without toxicity or high immune responses.

You can find a wide variety of tools commercially available for iPS cell reprogramming, including premade lentiviral and retroviral plasmids containing stem cell factors. For retroviral packaging, Platinum retroviral packaging cells exhibit longer stability and produce higher yields of retroviral structure proteins (gag, pol and env) compared to other cells, allowing for high-titer retroviral packaging with a single plasmid transfection. In addition, the well-published, passage-independent SNL feeder cells help support iPS cell culture.