June 19, 2017
How Does Sendai Virus Reprogram Cells?Learn more at http://www.lifetechnologies.com/cytotune
This video demonstrates how Sendai virus, found in the Cytotune®-iPS Sendai Reprogramming Kit, reprograms somatic cells to generate induced Pluripotent Stem Cells (iPSCs)
How Does Sendai Virus Reprogram Cells?
Induced pluripotent stem cells (iPSCs) are genetically reprogrammed somatic cells that exhibit a pluripotent stem cell state similar to embryonic stem cells. The discovery in 2006 that human and mouse fibroblasts could be reprogrammed to generate iPSCs with qualities remarkably similar to embryonic stem cells has created a valuable new source of pluripotent cells.
Fibroblasts, similar to other somatic cell types, do not express high levels of the transcription factors Oct4, Sox2, Klf4, and c-Myc under normal conditions. High levels of expression of these four genes will cause reprogramming of the fibroblast and it will become pluripotent.
There are multiple methods to generate iPSCs. Viruses such as retroviral vectors require integration of the viral genome into the host's chromosomes to express reprogramming genes. Normal gene transcription allows the inserted Oct4, Sox2, Klf4, and c-Myc transgenes to be expressed along with the host's genes. Integration of viral DNA into the host genome disrupts the genome of the cells. This alteration can render the iPSCs and their derivatives less safe for clinical application and can compromise compound screens or disease pathway analyses. On the other hand, Sendai virus is a single stranded negative-sense RNA virus that replicates in the cytoplasm. This means it does not integrate into the host's genome.
Sendai virus replicates independent of cell cycle, unlike other approaches where the exogenous genes are expressed only as the cell divides. Using this strategy, Sendai virus produces very high copy numbers of the target gene. The Cytotune® iPS Sendai Reprogramming Kit contains four Sendai virus-based reprogramming vectors, each expressing one of the four Yamanaka factors Oct4, Sox2, Klf4, and c-Myc.
These viral vectors are added to the dish of cells to be reprogrammed, incubated overnight, and the reprogramming process begins. After reprogramming, quantitative PCR testing shows that the Sendai virus does not remain in the iPSCs, allowing you to perform your research with iPSCs that have no genomic integration or viral remnants. Sendai virus is particularly useful when reprogramming patient derived blood cells, such as CD34 positive cells, T-cells, and PBMCs.
With its non-integrating capabilities, the Sendai virus within the Cytotune®-iPS Sendai Reprogramming Kit allows the use of iPSCs and their derivatives in a broad range of research experiments.