The modern human aryl hydrocarbon receptor is more active when ancestralized by genome editing.

The aryl hydrocarbon receptor (AHR) is a transcription factor that has many functions in mammals. Its best known function is that it binds aromatic hydrocarbons and induces the expression of cytochrome P450 genes, which encode enzymes that metabolize aromatic hydrocarbons and other substrates. All present-day humans carry an amino acid substitution at position 381 in the AHR that occurred after the divergence of modern humans from Neandertals and Denisovans.

Efficient high-precision homology-directed repair-dependent genome editing by HDRobust.

Homology-directed repair (HDR), a method for repair of DNA double-stranded breaks can be leveraged for the precise introduction of mutations supplied by synthetic DNA donors, but remains limited by low efficiency and off-target effects. In this study, we report HDRobust, a high-precision method that, via the combined transient inhibition of nonhomologous end joining and microhomology-mediated end joining, resulted in the induction of point mutations by HDR in up to 93% (median 60%, s.e.

Detection of unintended on-target effects in CRISPR genome editing by DNA donors carrying diagnostic substitutions.

CRISPR nucleases can introduce double-stranded DNA breaks in genomes at positions specified by guide RNAs. When repaired by the cell, this may result in the introduction of insertions and deletions or nucleotide substitutions provided by exogenous DNA donors. However, cellular repair can also result in unintended on-target effects, primarily larger deletions and loss of heterozygosity due to gene conversion.

Improved gRNA secondary structures allow editing of target sites resistant to CRISPR-Cas9 cleavage.

The first step in CRISPR-Cas9-mediated genome editing is the cleavage of target DNA sequences that are complementary to so-called spacer sequences in CRISPR guide RNAs (gRNAs). However, some DNA sequences are refractory to CRISPR-Cas9 cleavage, which is at least in part due to gRNA misfolding. To overcome this problem, we have engineered gRNAs with highly stable hairpins in their constant parts and further enhanced their stability by chemical modifications.

Comment on "Reintroduction of the archaic variant of in cortical organoids alters neurodevelopment".

Trujillo . (Research Articles, 12 February 2021, eaax2537) conclude that the reintroduction of an ancestral amino acid substitution in the protein NOVA1 drastically alters the development of brain organoids. We show that cell lines used by the authors carry heterozygous deletions of the target DNA sequence, providing another plausible explanation for the effects observed.