Here, in this chapter, we describe a detailed protocol for the method named Cas9-mediated protein evolution reaction or short CasPER. CasPER is based on the generation of large 300-600-bp mutagenized linear DNA fragments by error-prone PCR which are used as a donor for repair of double-strand break …

CRISPR-Cas9基因编辑技术就是通过人工设计的 sgRNA（guide RNA）来识别目的基因组序列，并引导 Cas9 蛋白酶进行有效切割 DNA 双链，形成双链断裂，损伤后修复会造成基因敲除或敲入等，最终达到对基因组DNA 进行修饰的目的。

CRISPR/Cas9 is a gene-editing technology which involves two essential components: a guide RNA to match a desired target gene, and Cas9 (CRISPR-associated protein 9)—an endonuclease which causes a double-stranded DNA break, allowing modifications to the genome (see figure 1).

CRISPR-Cas9 is a genome editing tool that is creating a buzz in the science world. It is faster, cheaper and more accurate than previous techniques of editing DNA and has a wide range of potential applications.

Cas9 is an accurate method of treating diseases due to the targeting of the Cas9 enzyme only affecting certain cell types. The cells undergoing the Cas9 therapy can also be removed and reintroduced to provide amplified effects of the therapy.

Jul 1, 2018 · Cas9 is an RNA-guided endonuclease that introduces double-stranded breaks (DSBs) at almost any target DNA locus where a protospacer-adjacent motif (PAM; NGG) is present, yielding several hundred thousand genomic target sites in an average eukaryotic genome (DiCarlo et al., 2013).

Jul 4, 2022 · Here, in this chapter, we describe a detailed protocol for the method named Cas9-mediated protein evolution reaction or short CasPER. CasPER is based on the generation of large 300–600-bp mutagenized linear DNA fragments by error-prone PCR which are used as a donor for repair of double-strand break mediated by Cas9 and subsequently integrated ...

The method employs error-prone PCR and Cas9-mediated genome integration of mutant libraries of large-sized donor variants into single or multiple genomic sites with efficiencies reaching 98-99%.

Jul 1, 2018 · Here we describe a method for robust directed evolution using mutagenesis of large sequence spaces in their genomic contexts. The method employs error-prone PCR and Cas9-mediated genome...

Aug 21, 2018 · The newly developed method is named CasPER and is building on existing technologies, such as CRISPR/Cas9, that has been used for genome engineering and re-programming in yeast during the last...