Research Techniques Made Simple: The Application of CRISPR-Cas9 and Genome Editing in Investigative Dermatology

Abstract

Designer nucleases have gained widespread attention for their ability to precisely modify genomic DNA in a programmable manner. These genome-editing nucleases make double-stranded breaks at specified loci, and desired changes can be made to modify, ablate, or excise target genes. This technology has been used widely to develop human disease models in laboratory animals and to study gene functions by silencing, activating, or modifying them. Furthermore, the recent discovery of a bacterially derived programmable nuclease termed clustered regularly interspaced palindromic repeats (CRISPR)-associated protein 9 (Cas9) has revolutionized the field because of its versatility and wide applicability. In this article, we discuss various modalities used to achieve genome editing with an emphasis on CRISPR-Cas9. We discuss genome-editing strategies to either repair or ablate target genes, with emphasis on their applications for investigating dermatological diseases. Additionally, we highlight preclinical studies showing the potential of genome editing as a therapy for congenital blistering diseases and as an antimicrobial agent, and we discuss limitations and future directions of this technology.

COL7A1 Editing via CRISPR/Cas9 in Recessive Dystrophic Epidermolysis Bullosa

“… 1 , 2 , 3 , 4 , 5 , 6 , 7 The versatility of CRISPR/Cas9, including simple design and the possibility of targeting multiple genes simultaneously, has aided the rapid evolution of this groundbreaking technology in recent years. 8 Directed by a specifically designed single guide RNA (sgRNA) to the respective DNA target site, the Cas9 nuclease performs DNA double-strand breaks (DSBs), leading to the activation of the cellular repair machinery to correct the DNA damage either by non-homologous end joining (NHEJ) or, in the presence of a homologous DNA donor template, by homology-directed repair (HDR). 9 …”

Research Techniques Made Simple: CRISPR Genetic Screens

“…CRISPR-based screens demonstrate improved versatility, efficacy, and lower off-target effects compared with approaches such as RNAi (Ford et al, 2019;Guitart et al, 2016;Schuster et al, 2019). For a comprehensive description of the fundamentals of CRISPR-mediated genome editing, we refer to a previous Research Techniques Made Simple article (Guitart et al, 2016). In brief, the bacterial Cas enzyme (usually Cas9) is guided to a genomic DNA target by a single guide RNA (sgRNA), an approximately 20nucleotide sequence that specifies the genomic target, such as a protein-coding gene.…”

“…In a typical cell cultureebased screen, systematic loss-of-function of a set of candidates is applied to identify elements contributing to a phenotype of interest. RNA interference (RNAi) and transposon-based technologies have been used successfully, but since their development, CRISPR/Cas-based tools have become a preferred method for genetic screens (Doench, 2018;Ford et al, 2019;Guitart et al, 2016;Schuster et al, 2019).…”

Precision therapy for psoriasis patients

“…Несмотря на многочисленные исследования в области генетической предрасположенности к развитию болезней, а также изучение технологии редактирования генома (CRISP и другие), профилактическая медицина еще только формируется [9]. В то же время уже сегодня тесты на выявление генетических отклонений плода обязательны всем беременным, а количество исследований для оценки риска развития различных заболеваний ежегодно увеличивается [10].…”