Realizing the promise of prime editing for the study and treatment of genetic disorders requires efficient methods for delivering prime editors (PEs) in vivo. Here we describe the identification of bottlenecks limiting adeno-associated virus (AAV)-mediat
Here we describe the identification of bottlenecks limiting adeno-associated virus (AAV)-mediated prime editing in vivo and the development of AAV-PE vectors with increased PE expression, prime editing guide RNA stability and modulation of DNA repair. The resulting dual-AAV systems, v1em and v3...
For H2A.X mRNA detection, RNA was separated on 1% agarose containing 5.5% v/v formaldehyde 36.5–38% in MOPS and transferred by capillarity in 10x SSC (1.5 M NaCl and 150 mM tri-sodium citrate). For U7 snRNA northern blots, RNA was separated on 8% polyacrylamide gels (60% v/v ...
We also tested the insertion activity of ShCAST using a synthetic single guide RNA (sgRNA) containing the 13-nt direct repeat from the delocalized crRNA (Strecker et al. 2019) by deleting different lengths of spacer sequence. We found that 16 nt is both sufficient and near the minimum ...
Each single-guide RNA (sgRNA), together with four individual constructs, was cotransfected into HEK293T cells. The editing efficiency was analyzed by Sanger sequencing, and the results showed that all 4 editors induced simultaneous A/C conversions. The mean efficiency of CABE-1 was higher than ...
Macbeth MR, Schubert HL, VanDemark AP, Lingam AT, Hill CP, Bass BL (2005) Inositol hexakisphosphate is bound in the ADAR2 core and required for RNA editing. Science 309:1534–1539 Article ADS Google Scholar Huang YH, Grasis JA, Miller AT, Xu R, Soonthornvacharin S, Andreotti AH, Tso...
To target PD-L1 for CRISPR/Cas9 gene editing, we first identified two single guide RNA (sgRNA) sequences located on PD-L1 exon 3. The first sgRNA recognizes the forward strand of human PD-L1 near the beginning of exon 3 that allows editing by Cas9 at approximately base pair 82 (g82)...
was used with 20 individual single guide RNAs in rice protoplasts to produce near-saturated mutagenesis (73.21%) for a 56-amino-acid portion of the rice acetyl-coenzyme A carboxylase (OsACC). We also applied STEME-1 and STEME-NG for directed evolution of theOsACCgene in rice and obtained ...
In this work, the space-time MORe DWR (Model Order Reduction with Dual-Weighted Residual error estimates) framework is extended and further developed for single-phase flow problems in porous media. Specifically, our problem statement is the Biot system w
Adenine transversion editors enable precise, efficient A•T-to-C•G base editing in mammalian cells and embryos Article15 June 2023 Chemical modifications of adenine base editor mRNA and guide RNA expand its application scope ArticleOpen access24 April 2020 ...