Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology has been widely used in gene editing by inducing gene mutations and activating or inhibiting gene expression. Guide RNA (gRNA) and CRISPR-associated (Cas) nuclease are the two essential components in the CRISPR/Cas system. Based on the structure and function of Cas proteins, CRISPR/Cas systems can be divided into class I and class II. The class I systems are composed of multi-subunit Cas protein complexes, and the class II systems utilize a single Cas protein. CRISPR/Cas-9 system has been widely used in genetic engineering due to its relatively simple structure. The Cas-9 protein is a large multi-region DNA endonuclease responsible for cleaving the target DNA to form a double-strand break. The Cas-9 protein is guided to a target genome locus by the gRNA, which consists of CRISPR RNA (crRNA) and trans-activating CRISPR RNA (tracrRNA). CRISPR/Cas9 technology has revolutionized large-scale functional genomic screening.
CRISPR libraries are designed to cause three types of gene expression modifications: knockout, suppression, and activation. CRISPR interference (CRISPRi) libraries are an alternative to knockout libraries. In contrast to knockout libraries, CRISPRi libraries use inactive Cas9 (dCas9) to inhibit transcription. The dCas9 protein can inhibit transcription by blocking RNA polymerase directly or by repressing transcription as a fusion protein to a repressive effector domain. Gain-of-function (GoF) screen using CRISPR-mediated gene activation (CRISPRa) technology is a different approach in gene candidate identification. CRISPRa technology utilizes a similar method of transcriptional regulation as CRISPRi technology to fuse the dCas9 protein to a known activated domain protein. However, the goal of CRISPRa technology is to increase the level of gene expression. In general, CRISPR libraries come in arrayed format and pooled format. In the arrayed format, each element of the library is compartmentalized, while in the pooled format, each element of the library is present in a single pool. Arrayed libraries are often used to study the effects of gene knockouts on complex phenotypes. Array libraries can also be used to study subtle phenotypes that require parallel analysis of thousands of cells, each of which has the same perturbation. The pooled libraries are relatively inexpensive to perform compared to the arrayed libraries, and rarely require specialized equipment. Most CRISPR libraries use lentiviruses for cell transduction. Adeno-associated viruses (AAVs), adenoviruses (AdVs) or retroviruses are also employed for gene delivery in some CRISPR libraries.
Amerigo Scientific offers high-quality pre-made CRISPR libraries for high-throughput screening of important molecular targets, including whole-genome CRISPR knockout libraries, whole-genome CRISPRa libraries, whole-genome CRISPRi libraries, and pathway-specific CRISPR knockout libraries. The libraries enable genetic knockout, activation, or repression for genome-wide and pathway-specific screening.
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