The genome editing revolution

Seventy years after deciphering the structure of DNA, we are experiencing a revolution in genome editing.Sequencing and synthesis of DNA is cheaper and faster than ever, leading to the rapid advancement of the fields of molecular biology and biotechnology.The development of DNA editing and engineering tools advanced our fundamental understanding of biology, which allowed us to develop societally relevant applications.CRISPR-Cas editing tools have revolutionized the field of genome editing due to their simplicity, accuracy, and efficiency across all forms of life. A series of spectacular scientific discoveries and technological advances in the second half of the 20th century have provided the basis for the ongoing genome editing revolution. The elucidation of structural and functional features of DNA and RNA was followed by pioneering studies on genome editing: Molecular biotechnology was born. Since then, four decades followed during which progress of scientific insights and technological methods continued at an overwhelming pace. Fundamental insights into microbial host-virus interactions led to the development of tools for genome editing using restriction enzymes or the revolutionary CRISPR-Cas technology. In this review, we provide a historical overview of milestones that led to the genome editing revolution and speculate about future trends in biotechnology. A series of spectacular scientific discoveries and technological advances in the second half of the 20th century have provided the basis for the ongoing genome editing revolution. The elucidation of structural and functional features of DNA and RNA was followed by pioneering studies on genome editing: Molecular biotechnology was born. Since then, four decades followed during which progress of scientific insights and technological methods continued at an overwhelming pace. Fundamental insights into microbial host-virus interactions led to the development of tools for genome editing using restriction enzymes or the revolutionary CRISPR-Cas technology. In this review, we provide a historical overview of milestones that led to the genome editing revolution and speculate about future trends in biotechnology. The development of molecular biology is based on a number of groundbreaking discoveries excellently reviewed in ‘The Eighth Day of Creation’ [1.Judson H.F. Gratzer W. The eighth day of creation.Nature. 1997; 386: 344Google Scholar] (Table 1). It all started with the identification of DNA as the storage polymer for genetic information [2.Avery O. et al.Studies on the chemical nature of the substance causing transformation of the pneumococcal types. Induction by a desoxyribonucleic acid fraction isolated from pneumococcus type III.J. Exp. Med. 1944; 79: 137-158Crossref PubMed Google Scholar]. 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Table 1 shows major discoveries in molecular biology, from establishing the Central Dogma to state-of-the-art genome editing and high-throughput analysis.Table 1Milestones in molecular biologyYearMilestones in molecular biologyKey players1944DNA carrier of genetic informationAvery, Macload and McCarty1953DNA structureWatsonaNobel laureate. and CrickaNobel laureate., Franklin and WilkinsaNobel laureate.1954Protein sequencingSangeraNobel laureate.1958Central DogmaCrick1959Protein structureKendrewaNobel laureate. and PerutzaNobel laureate.1956–1960DNA and RNA polymerase and ribosomeA. KornbergaNobel laureate., Ocheaa,Nobel laureate. and PaladeaNobel laureate.1962Gene expression/regulationJacobaNobel laureate. and MonodaNobel laureate.1966Genetic codeNirenbergaNobel laureate., KhoronaaNobel laureate., HolleyaNobel laureate.1972Restriction enzymeArberaNobel laureate., SmithaNobel laureate., NathansaNobel laureate.1972DNA recombinationBergaNobel laureate.1975DNA sequencingSangeraNobel laureate., MaximaNobel laureate.1986PCR and site-directed mutagenesisMullisaNobel laureate. and SmithaNobel laureate.1992Laboratory evolutionStemmer and ArnoldaNobel laureate.1997RecombineeringSteward and Murphy1995Genome sequencingCollins, Lander, and Venter1998RNAiFireaNobel laureate. and MelloaNobel laureate.2000Ribosome structure and functionRamakrishnanaNobel laureate., SteitzaNobel laureate., YonathaNobel laureate.2001RNA polymeraseR. KornbergaNobel laureate.2005–2022CRISPR-CasbFor CRISPR milestones, see Table 3.CharpentieraNobel laureate. and DoudnaaNobel laureate.2020–2022Protein structure prediction(e.g., Alphafold2, ESMFold, RoseTTAFold)DeepMind (Google), ESM (Meta), Baker lab (University of Washington)a Nobel laureate.b For CRISPR milestones, see Table 3. Open table in a new tab In parallel to the aforementioned fundamental discoveries on the storage and processing of genetic information, pioneering genetic studies were performed on bacteria and/or bacterial viruses (bacteriophages). This has resulted in unraveling many basic genetic principles, including gene expression and control thereof (e.g., the lac operon of Escherichia coli [13.Jacob F. Monod J. Genetic regulatory mechanisms in the synthesis of proteins.J. Mol. Biol. 1961; 3: 318-356Crossref PubMed Google Scholar]), but also in revealing a wide range of bacterial defense systems [14.Luria S.E. Delbrück M. Mutations of bacteria from virus sensitivity to virus resistance.Genetics. 1943; 28: 491Crossref PubMed Google Scholar,15.Arber W. Linn S. DNA modification and restriction.Annu. Rev. Biochem. 1969; 38: 467-500Crossref PubMed Google Scholar] as well as phage attack strategies [16.Lwoff A. Lysogeny.Bacteriol. Rev. 1953; 17: 269-337Crossref PubMed Google Scholar] (Table 1). Altogether, this fundamental research led to the discovery of enzymes with the potential for genetic engineering, such as specific DNA endonucleases [15.Arber W. Linn S. DNA modification and restriction.Annu. Rev. Biochem. 1969; 38: 467-500Crossref PubMed Google Scholar,17.Nathans D. Smith H.O. Restriction endonucleases in the analysis and restructuring of DNA molecules.Annu. Rev. Biochem. 1975; 44: 273-293Crossref PubMed Google Scholar] (Table 1 and Figure 2A ). Combining specific type II restriction nucleases with DNA ligase allowed ‘cut-and-paste’ engineering of DNA fragments of a primate virus, simian virus 40 (SV40) [18.Mertz J.E. Davis R.W. of DNA by restriction S. A. PubMed Scopus (0) Google Scholar]. spectacular was an in which a of enzymes a polymerase, a DNA polymerase, and a DNA allowed the of a DNA from a into the genome et for new genetic information into DNA of simian virus DNA phage genes and the operon of Escherichia S. A. PubMed Scopus Google Scholar]. an Escherichia coli was as a for a DNA a gene from transformation to coli the gene was the coli to on bacterial in and expression of genes in Escherichia S. A. PubMed Google Scholar]. In a the was to genes from a in coli et and transcription of DNA in Escherichia S. A. 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