11 research outputs found
A Young Seedling Stripe2 phenotype in rice is caused by mutation of a chloroplast-localized nucleoside diphosphate kinase 2 required for chloroplast biogenesis
Natural variation in expression of genes associated with carotenoid biosynthesis and accumulation in cassava (Manihot esculenta Crantz) storage root
Chloroplast FtsZ assembles into a contractible ring via tubulin-like heteropolymerization
Highly parallel genome variant engineering with CRISPR–Cas9
Understanding the functional effects of DNA sequence variants is of critical importance for studies of basic biology, evolution, and medical genetics; however, measuring these effects in a high-throughput manner is a major challenge. One promising avenue is precise editing with the CRISPR-Cas9 system, which allows for generation of DNA double-strand breaks (DSBs) at genomic sites matching the targeting sequence of a guide RNA (gRNA). Recent studies have used CRISPR libraries to generate many frameshift mutations genome wide through faulty repair of CRISPR-directed breaks by nonhomologous end joining (NHEJ) 1 . Here, we developed a CRISPR-library-based approach for highly efficient and precise genome-wide variant engineering. We used our method to examine the functional consequences of premature-termination codons (PTCs) at different locations within all annotated essential genes in yeast. We found that most PTCs were highly deleterious unless they occurred close to the 3' end of the gene and did not affect an annotated protein domain. Unexpectedly, we discovered that some putatively essential genes are dispensable, whereas others have large dispensable regions. This approach can be used to profile the effects of large classes of variants in a high-throughput manner