Researchers from the Institute of Biophysics of the Chinese Academy of Sciences achieved conditional targeted genome editing in C. elegans by using somatic cells to express transcription activator-like effector nucleases (TALENs). The research results were published online in the August 18 issue of "Nature Biotechnology" (Nature Biotechnology) magazine.

Researcher Guangshuo Ou and Dr. Wei Li of the Institute of Biophysics, Chinese Academy of Sciences are co-corresponding authors of this paper. Researcher Ou Guangshuo's main research direction is to develop in vivo fluorescence sequential imaging technology combined with biochemistry and genetics, using nematodes as a model to study how the cytoskeleton and signal transduction proteins regulate the development of the nervous system.

For decades, nematodes have become a universal model system for studying basic biology and human diseases, and many genetic tools and resources are suitable for this organism. Of the 20,377 predicted protein-encoding genes, 6,764 genes currently achieve gene knockout or null mutation mainly through random chemical mutagenesis or targeted gene knockout based on the Mos1 transposon. However, the current technology cannot conditionally edit the wild-type nematode genome in a targeted manner, limiting scientists' answers to biological questions.

TALEN technology is a new genome editing technology developed by scientists in recent years, which can generate site-specific mutations in the genome. TALENs are artificially engineered restriction enzymes composed of a non-specific endonuclease cleavage domain (FokI) fused with a custom repeat domain that recognizes a predictable DNA sequence. The specific DNA recognition domain is composed of some very conservative repeating amino acid sequence modules, each module is generally composed of 34 amino acids (aa), in which the amino acid type of the 12th and 13th positions is variable and determines the recognition target of the module Point specificity. By binding the DNA recognition domain to the target site, the cleavage domain of FokI forms a dimer, which can specifically induce DNA double-strand breaks at the targeted site. After the damaged DNA is repaired at the non-homologous end junction, it will Due to the random increase or decrease of bases, the breakpoint mutagenesis is deleted or inserted.

Although scientists have applied TALENs to edit the genomes of a wide range of organisms, they are still limited to nematode cultured cells or embryos, and germ cell lines. So far it is unclear whether TALENs can be applied to somatic cells of multicellular organisms.

In this article, the researchers developed a method to achieve conditional gene knockout by expressing TALENs in C. elegans somatic cells. By using inducible or tissue-specific promoters to control the plasmids encoding TALENs to transform germ cells, the researchers also observed effective genetic modification in specific developmental stages and tissues, and the resulting phenotype. The Cor-1 gene is responsible for encoding homologues of human severe combined immunodeficiency (SCIED) protein coronin. In further research, the researchers used this method to bypass the embryo's need for cor-1, and determined that cor-1 plays a crucial role in cell migration in the larval Q-cell line.

These findings indicate that the expression of TALENs in model organism cells is a versatile tool suitable for functional genomics research.

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