Brief intro:
- Author: Feiyu Zhao, Tao Zhang, Xiaodi Sun, Xiyun Zhang, Letong Chen, Hejun Wang, Jinze Li, Peng Fan, Liangxue Lai,Tingting Sui and Zhanjun Li
- Journal: Nat Commun
- Doi: https://www.doi.org/10.1038/s41467-023-41320-8
- Publication Date: 2023 Sep 8
Products/Services used in the paper
Quotation shows PackGene:AAV injection PackGene Biotech (China) created the AAV-mini-RfxCas13d-Pcsk9-crRNA2 expression cassettes and produced the viruses (https://www.packgene.cn/about-us/). An AAV injection assay was performed as previously described44. Production of AAV8-RfxCas13d vectors was performed by PackGene Biotech (China).The AAV-mini-RfxCas13d-Pcsk9-crRNA2 titer was 2 × 1012 genome copies (GCs)/mL.
Research Field:Liver, blood
AAV Serotype:AAV8
Targeted organ:Liver, blood
Animal or cell line strain:mouse
Abstract
The small size of the Cas nuclease fused with various effector domains enables a broad range of function. Although there are several ways of reducing the size of the Cas nuclease complex, no efficient or generalizable method has been demonstrated to achieve protein miniaturization. In this study, we establish an Interaction, Dynamics and Conservation (IDC) strategy for protein miniaturization and generate five compact variants of Cas13 with full RNA binding and cleavage activity comparable the wild-type enzymes based on a combination of IDC strategy and AlphaFold2. In addition, we construct an RNA base editor, mini-Vx, and a single AAV (adeno-associated virus) carrying a mini-RfxCas13d and crRNA expression cassette, which individually shows efficient conversion rate and RNA-knockdown activity. In summary, these findings highlight a feasible strategy for generating downsized CRISPR/Cas13 systems based on structure predicted by AlphaFold2, enabling targeted degradation of RNAs and RNA editing for basic research and therapeutic applications.
About PackGene
PackGene Biotech is a world-leading CRO and CDMO, excelling in AAV vectors, mRNA, plasmid DNA, and lentiviral vector solutions. Our comprehensive offerings span from vector design and construction to AAV, lentivirus, and mRNA services. With a sharp focus on early-stage drug discovery, preclinical development, and cell and gene therapy trials, we deliver cost-effective, dependable, and scalable production solutions. Leveraging our groundbreaking π-alpha 293 AAV high-yield platform, we amplify AAV production by up to 10-fold, yielding up to 1e+17vg per batch to meet diverse commercial and clinical project needs. Moreover, our tailored mRNA and LNP products and services cater to every stage of drug and vaccine development, from research to GMP production, providing a seamless, end-to-end solution.
