siRNA In Vitro Screening Service
In order to identify and validate the targets of interest, researchers use siRNA collections with readouts to take full advantage of the potential of siRNAs present for drug discovery. Creative Biolabs now provides convenient siRNA in vitro screening services. Adding siRNA screening to your early drug discovery screening programs enables customers to identify targets that traditional biochemical and cell-based screening & profiling may miss. Our sophisticated technologies and professional scientists are available to provide you with customized siRNA screening services.
siRNA In Vitro Screening Introduction
The whole process of siRNA in vitro screening service is shown in Figure 1. Creative Biolabs provides appropriate scale of RNAi library to screen (e.g., genome wide, kinase family or customer-defined) and develops robust and reproducible assays for identification and selection of candidate genes. Meanwhile, Creative Biolabs will choose the appropriate type of RNAi library, including chemically synthesized siRNAs, short hairpin siRNA (shRNA) and endoribonuclease-derived siRNAs (esiRNA), based on your genes.
Figure 1. Flowchart of RNAi high-throughput screening. (Gao, 2014)
There are two formats of siRNA in vitro screening services: a pooled screening format, in which the library is introduced randomly into cells (Figure 2a), or an arrayed screening format, in which single genes are targeted by reagents in individual wells of a microtiter plate (Figure 2b).
Figure 2. Two formats of siRNA in vitro screening. (Mohr, 2010)
Arrayed Screening
- Purposes: This screening way is suitable for high-content screening assays and suitable for primary cells and neurons. Also, custom-made libraries are available. Moreover, the arrayed screening can analyze detections of responses to an external stimulus (e.g., stress, drug treatment, pathogen, signaling ligand, or metabolic substrate); and observe multiple, related phenotypes can be assayed in a single screen.
- Tools: In this screening, we will utilize three types of tools. The first one is chemically synthetic siRNA. It is suitable for short-term gene silencing. The second one is the vector-encoded shRNA. It is suitable for long-term and stable gene silencing. The third one is for special conditions, and we provide lentiviral-based shRNA libraries for primary cells, that are difficult to transfect.
- Methods: To observe the readouts, there are two methods we provide. Depending on different conditions, detection of the assay is done via measuring colorimetric, fluorescence, or luminescent readouts at the total well level (plate-reader screens). The other method is to measure fluorescent readouts at the cellular or subcellular level using imaging.
Pooled Screening
- Purposes: This screening way is suitable for cell viability/proliferation assays to identify drug resistance genes. This way is also for mammalian cells by a viral-encoded short hairpin RNA (shRNA). Besides that, pooled screening could be used for genome-wide RNA. Different purposes have their own suitable ways.
- Tools: In pooled screening, the main tool is viral-encoded shRNA.
- Methods: This is important that using pooled screening to select should be careful about the phenotype. Pooled screening is based on phenotype being studied. Customers and researchers should know the phenotype before deciding to use this way for selecting the targets. For pooled screening, the selections can be divided into two types: positive screens and negative screens, shown in Figure 3. In terms of positive selection screens, it utilizes fluorescence-activated cell sorter (FACS). During the positive screens, it makes most cells to die or do not pass the selection and then sort "win" cells. Then, sequencing "win" cells to find out which one is expected. Conversely, negative selection screens make most cells to survive and then perform Next Generation Sequencing (NGS) on a control sample (no selection).
Figure 3. The processes of the positive screen and negative screen.
Creative Biolabs aspires to become the trusted solution provider of your first choice. Our advanced technologies and highly experienced staffs are committed to advancing your program with our siRNA in vitro screening services and reducing the overall development timeline. If you have any special requirements in the siRNA screening services, please don't hesitate to contact us.
References
- Gao, S.; et al. (2014). Applications of RNA interference high-throughput screening technology in cancer biology and virology. Protein & Cell. 5(11): 805-815.
- Mohr, S.; et al. (2010). Genomic screening with RNAi: results and challenges. Annual review of biochemistry. 79: 37-64.
