Today, researchers realizes the importance of knowing the events that occur at molecular level within a cell, the building block of a living organism. Many cellular events occur at molecular and sub-molecular level. Investigative efforts to understand these processes are tedious and time consuming. Before reaching any conclusion, scientists need to check their findings several times. As a helpful resource for life scientists there are number of commercially available research services offered by contract research laboratories (biology CRO). Developing synthetic RNA for RNA interference (RNAi) systems is one such instance of custom services.
RNA interference (RNAi) is a potent tool that life scientists frequently use to discover the structural and functional marvels at cellular level. Researchers can insert foreign DNA or RNA sequences in host cells, the process referred to as transfection. Once a cell is transfected, the exogenous RNA intercepts the host cell’s mRNA before it can translate its specific protein. In effect, RNAi suppresses gene expression, a phenomenon known as gene silencing. Though the process sounds simple, it is in realty far away from being that. Scientists come across a multitude of problems when trying to set up RNAi systems. To begin with, DNA encoding gene of interest (or siRNA/shRNA targeting protein of interest) should be designed, synthesized (or cloned) and transfected into host cells, which necessitates the use of transfection reagents. The effect of transfection reagents too varies from cell to cell. Some transfection reagents work like a charm with certain types of cells but may show poor transfection efficiency when used with different cell lines. Some cell lines can be readily transfected while others may be transfection resistant not to mention the changes they undergo in culture medium especially after several passages.
Transfection itself may many a times be transient, implying that the exogenous RNA fails to become a part of the transfected cell’s genome. The desired results therefore happen for a short duration of time, not enough for in-depth studies. To enable life scientists to conduct detailed investigations, foreign RNA must be internalized (and integrated into genomic DNA) in host cell nucleus, so it can be expressed in all daughter cells.
Cytotoxicity is another issue affecting the success of transfection systems. In a nutshell, life scientists need to make a host of careful decisions about diverse aspects of a viable RNAi system. This way they lose precious time in optimizing rather than concentrating on exploring their topic of interest. Custom CRO services can be utilized to save valuable research time and resources.
RNAi custom services typically offer to design siRNA or shRNA-encoding plasmid genetic vector to target protein of interest. This may include pool of different siRNA (usually 3 – 5) targeting different parts of mRNA. siRNAs are synthetically produced and tested for gene silencing efficiency in vitro. High-throughput approaches include siRNA screening of specific gene family or genome-wide siRNA screens. Gene silencing effect is measured using percent of mRNA knockdown experimentally measured via qPCR (RT-PCR method). Alternative readout is reduction of protein expression levels, but automatic quantitative systems are relatively rare and automatic protein expression contract research services are offered only by Western Blot Inc in USA.
The custom RNAi services may involve formulation of effective in vitro transfection reagents, designing and development of targeted in vivo delivery systems, adequate screening and optimization of all parameters of the RNAi system. Custom RNAi services offered by Altogen Labs are cost effective and save scientists the time-consuming setting up their own approaches to develop functional and effective small RNAs.