Solution for Library Construction and Screening of Target Genes    

Cells use intricate gene networks to adapt to a variety of situations and surroundings. High-throughput genome-wide screening is made possible by CRISPR technology, which makes it easier to identify gene targets associated with certain phenotypes or to customize cellular function. Prior to the development of CRISPR technology, RNA interference (RNAi)-based screening was extensively used; however, because of poor gene suppression and off-target effects on mRNA, it was difficult to correlate phenotypes with gene knockdown.

Utilizing CRISPR-Cas Screening

Using a gRNA library for CRISPR-Cas screening has shown to be an effective method for identifying the genes, pathways, and processes linked to certain biological features or phenotypes. The identification of genes associated with bacterial invasion and resistance to chemicals or antibiotics has been made possible through the use of CRISPR-Cas9 screening. However, because CRISPR knockdown (CRISPR-KO) screening results in irreversible gene disruption, it is not able to reversibly modulate gene expression.

CRISPRi/a Screening

CRISPRi/a was created to overcome this restriction by allowing for the reversible expression of genes and the tweaking of their levels for screening purposes. A useful synthetic biology method that clarifies the connection between genotype and phenotype and identifies engineering targets in intricate cellular networks is genome-wide CRISPRi/a screening. There are two common ways to perform CRISPRi screening: arrayed and pooled methods.

Figure 1. Pooled and Arrayed CRISPR Screening (A) Pooled screening: cells are treated to a combination of Cas9 vector and sgRNA library delivery. High-throughput facilities use sgRNA sequencing to identify target genes and screen cells with certain characteristics. (B) Arrayed Screening: Individual cells are treated to independent delivery of the Cas9 and sgRNA vectors. The production of the chosen cells with the required characteristics is examined. (Jeong SH, et al., 2023)

Pooled CRISPR Screening

Target genes in pooled CRISPR screening are found by sequencing the sgRNA isolated from cells exhibiting certain phenotypic changes following the introduction of a large sgRNA library. For example, an E. coli sgRNA library containing around 60,000 sgRNAs was used to screen for genes relevant to auxotrophic response and antibiotic resistance.

Arrayed CRISPRi Screening

One sgRNA targeting one gene per well is inserted into cells in a multi-well plate during arrayed CRISPRi screening, and the phenotype that appears in each cell is monitored. Using a sgRNA library that targets putative L-proline transporter genes in Corynebacterium glutamicum, for instance, an L-proline exporter for L-proline hyperproduction was found.

Comprehensive CRISPR Library Screening Services of Target Genes

Creative Biogene's CRISPR/Cas9 Platform^CB stands at the forefront of the genome editing field, offering an extensive range of CRISPR library screening services. From screen design and cell line selection to the bioinformatics analysis of screen results, our services cover the entire spectrum of genome editing needs.

Specialized Screening Platforms

• CRISPR Knockout Screening: Identify essential genes and uncover new roles using CRISPR knockout screening. This approach efficiently identifies potential drug targets and facilitates drug development.
• CRISPRi Screening: Investigates the function of genes by reducing their expression without achieving a total knockout. CRISPRi screening is a great way to evaluate gene function in critical genes that become lethal when deleted.
• CRISPRa Screening: Use CRISPR activation to increase gene expression and investigate gain-of-function effects. This method sheds light on how genes function in their native environments.

Advantages of Our Services

• High-Throughput Capabilities: Our screening services are equipped with high-throughput capabilities that facilitate the swift identification of target genes that are crucial for a range of biological processes.
• Diverse Functional Assays: To thoroughly assess gene function, we provide a variety of functional assays, such as in vivo and cell-based assays.
• Ready-to-Use CRISPR Libraries: Our platform provides multiple CRISPR libraries ready for immediate use, streamlining the screening process and saving time and resources.
• Expert Support and Trained Staff: Every step of the screening process, from data analysis to experimental design, is carried out with the utmost care by our team of skilled experts and trained personnel.

Related Services and Products

Services

CRISPR Library Screening Service
Custom CRISPRi/CRISPRa Library Services
Base Editing Library Screening Service

Products

CRISPR Lentiviral Library
CRISPR gRNA Screening Kit