Correction of β-thalassemia mutant by base editor in human embryos

Abstract

β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB −28 (A>G) mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor (BE) system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB −28 (A>G) mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB −28 (A>G) homozygous mutation. Data showed that base editor could precisely correct HBB −28 (A>G) mutation in the patient’s primary cells. To model homozygous mutation disease embryos, we constructed nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured (IVM) oocytes. Notably, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB −28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.Electronic supplementary materialThe online version of this article (doi:10.1007/s13238-017-0475-6) contains supplementary material, which is available to authorized users.

Base pair editing in goat: nonsense codon introgression into FGF5 results in longer hair

“…On a genome‐wide scale using high coverage whole genome resequencing data for five mutant animals (> 40 × ) and four controls, one additional off‐target mutation predicted at four different target sites in all the animals was revealed, which was confirmed to be somatic (Figs A,B). Along with previous studies in human embryos , mice , and our recently study in sheep , these results indicated a low incidence of BE3‐induced off‐target mutagenesis, and the off‐target mutations were dependent largely on sgRNA design.…”

Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage

“…Together, these components enable efficient and permanent C•G to T•A base pair conversion in bacteria, yeast 4,9 , plants 10,11 , zebrafish 8,12 , mammalian cells 3–8,13,14 , mice 8,15,16 , and even human embryos 17,18 . Base editing capabilities have expanded through the development of base editors with different protospacer-adjacent motif (PAM) compatibilities 7 , narrowed editing windows 7 , enhanced DNA specificity 8 , and small-molecule dependence 19 .…”

Non-Structural Carbohydrates Accumulation in Contrasting Rice Genotypes Subjected to High Night Temperatures

“…Summarizing, rice plants can be affected by complex genome x environment x management interactions which results in phenotypic plasticity as a result of the variability of genetic components. Whether by on side, recently advances have been made in genetic analysis, propitiated by development of new approaches as such CRISPR/Cas (Cong et al, 2013) and base editor (Liang et al, 2017), on the other hand, in respect to non-invasive physiological phenotyping, there are scarcity of large-scale analyses of the underlying physiological mechanisms that plants trigger in response to environmental stimulus. In this sense, even in an initial phase, our data indicate that external phenotype such as spikelet sterility and 1000-grain weight is determined by the sum of the complex interactions resultant of an internal, physiological, and biochemical phenotype.…”