Plant Genetic Modification by CRISPR/CAS9

Lifeasible has developed a transgenic plant platform based on CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR associated protein9) to provide engineered plants with enhanced properties such as high yields and nutrients, resistance to disease, pests, or drought tolerance.

CRISPR/Cas9 gene editing is a technique that rapidly edits genomic DNA at a very high efficiency and specificity following the same mechanism used by prokaryote to defense viruses or other foreign DNA invasions. Cas9 is an RNA -guided DNA endonuclease enzyme associated with the CRISPR. sgRNA (single-guide RNA), which packages with multiple crRNAs and the tracrRNA, can combine Cas9 and guide this complex to a complementary sequence in the target DNA. Then the HNH and RuvC domains of Cas9 cut DNA on 3bp upstream of the PAM (protospacer adjacent motifs) site (typically NGG), making DSBs (DNA double strand breaks). The DSBs are repaired by either NHEJ (non-homologous end joining) or HDR (homology directed repair) allowing for nucleotide insertions, deletions, substitutions or site-specific mutations in the broken regions. The simple design of sgRNA strictly follows Watson-Crick base pairing and has made CRISPR/Cas9 a very effective and promising gene editing tool.

Figure 1. CRISPR/Cas9-mediated genomic modifications (only nucleotide insertion is shown here)

Lifeasible has a specialized molecular biology laboratory with the cutting-edge instrument. Our experts of the cell and molecular biology are devoted to providing industry-leading custom sgRNA design and vector construction services to customers all over the world. We can offer single and multiple genes knock-down, knock-out or knock-in, site-specific mutation even single nucleotide mutation according to alternative DNA repair pathways. We also can perform high throughput screening by the sgRNA library. With fully-fledged transgenic technologies in many species such as Arabidopsis, rice, wheat, alfalfa, corn, cotton, sorghum, soybean, sugarbeet, cabbage, tobacco, eggplant, cucumber, potato, petunia, oilseed rape, we are equipped to serve the diverse requirements of our customers. Lifeasible also has advanced CRISPR/Cpf1 system for more efficient and specific gene editing. The system has many advantages:

• Requiring only a crRNA for successful targeting.
• Causing a 'staggered' cut end.
• Increasing opportunity of repeated rounds of DNA cleavage by cleaving DNA 18-23 bp downstream from the PAM site.
• Providing alternate target sites in T rich regions.

Experimental process:

For more information please contact Lifeasible.