Gene Repair of iPSC Line with GARS (G294R) Mutation of CMT2D Disease by CRISPR/Cas9

Objective Charcot-Marie-Tooth disease(CMT)severely affects patient activity,and may cause disability.However,no clinical treatment is available to reverse the disease course.The combination of CRISPR/Cas9 and iPSCs may have therapeutic potential against nervous diseases,such as CMT.Methods In the present study,the skin fibroblasts of CMT type 2D(CMT2D)patients with the c.880G>A heterozygous nucleotide mutation in the GARS gene were reprogrammed into iPSCs using three plasmids(pCXLE-hSK,pCXLE-hUL and pCXLE-hOCT3/4-shp5-F).Then,CRISPR/Cas9 technology was used to repair the mutated gene sites at the iPSC level.Results An iPSC line derived from the GARS(G294R)family with fibular atrophy was successfully induced,and the mutated gene loci were repaired at the iPSC level using CRISPR/Cas9 technology.These findings lay the foundation for future research on drug screening and cell therapy.Conclusion iPSCs can differentiate into different cell types,and originate from autologous cells.Therefore,they are promising for the development of autologous cell therapies for degenerative diseases.The combination of CRISPR/Cas9 and iPSCs may open a new avenue for the treatment of nervous diseases,such as CMT.

Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

African swine fever virus(ASFV)is a lethal pathogen that causes severe threats to the global swine industry and it has already had catastrophic socio-economic effects.To date,no licensed prophylactic vaccine exists.Limited knowledge exists about the major immunogens of ASFV and the epitope mapping of the key antigens.As such,there is a considerable requirement to understand the functional monoclonal antibodies(mAbs)and the epitope mapping may be of utmost importance in our understanding of immune responses and designing improved vaccines,therapeutics,and diagnostics.In this study,we generated an ASFV antibody phage-display library from ASFV convalescent swine PBMCs,further screened a specific ASFV major capsid protein(p72)single-chain antibody and fused with an IgG Fc fragment(scFv-83-Fc),which is a specific recognition antibody against ASFV Pig/HLJ/2018 strain.Using the scFv-83-Fc mAb,we selected a conserved epitope peptide(221MTGYKH226)of p72 retrieved from a phage-displayed random peptide library.Moreover,flow cytometry and cell uptake experiments demonstrated that the epitope peptide can significantly promote BMDCs maturation in vitro and could be effectively uptaken by DCs,which indicated its potential application in vaccine and diagnostic reagent development.Overall,this study provided a valuable platform for identifying targets for ASFV vaccine development,as well as to facilitate the optimization design of subunit vaccine and diagnostic reagents.

A duplex TaqMan probe-based real-time PCR method for rapid detection and differentiation of the classical and VB strains of human immunodeficiency virus

Dear Editor,Human immunodeficiency virus(HIV)is a lentivirus that,if left untreated,can lead to acquired immunodeficiency syndrome(AIDS).The virus can be divided into two types based on genetic differences:HIV-1 and HIV-2.HIV-1 is widely distributed worldwide and is the primary strain responsible for the global AIDS epidemic due to its high replication capacity,greater likelihood of transmission,and more severe clinical symptoms(Oeschger et al.,2021).

The multi-kingdom microbiome catalog of the chicken gastrointestinal tract

Chicken is an important food animal worldwide and plays an important role in human life by providing meat and eggs.Despite recent significant advances in gut microbiome studies,a comprehensive study of chicken gut bacterial,archaeal,and viral genomes remains unavailable.In this study,we constructed a chicken multi-kingdom microbiome catalog(CMKMC),including 18,201 bacterial,225 archaeal,and 33,411 viral genomes,and annotated over 6,076,006 protein-coding genes by integrating 135 chicken gut metagenomes and publicly available metagenome-assembled genomes(MAGs)from ten countries.We found that 812 and 240 MAGs in our dataset were putative novel species and genera,respectively,far beyond what was previously reported.The newly unclassified MAGs were predominant in Phyla Firmicutes_A(n=263),followed by Firmicutes(n=126),Bacteroidota(n=121),and Proteobacteria(n=87).Most of the classified species-level viral oper-ational taxonomic units belong to Caudovirales.Approximately,63.24%of chicken gut viromes are predicted to infect two or more hosts,including complete circular viruses.Moreover,we found that diverse auxiliary metabolic genes and antibiotic resistance genes were carried by viruses.Together,our CMKMC provides the largest integrated MAGs and viral genomes from the chicken gut to date,functional insights into the chicken gastrointestinal tract microbiota,and paves the way for microbial interventions for better chicken health and productivity.