Precision Fluorescent Labeling of an Adeno-Associated Virus Vector to Monitor the Viral Infection Pathway.

Adeno-associated virus 2 (AAV2) is a common vehicle for the delivery of a variety of therapeutic genes. A better understanding of the process of infection of AAV2 will advance our knowledge of AAV2 biology and allow for the optimization of AAV2 capsids with favorable transduction profiles. However, the precise fluorescent labeling of an AAV2 vector for probing virus tracking without affecting the nature of the virus remains a challenge.

Site-Specific PEGylated Adeno-Associated Viruses with Increased Serum Stability and Reduced Immunogenicity.

Adeno-associated virus (AAV) is one of the most extensively studied and utilized viral vectors in clinical gene transfer research. However, the serum instability and immunogenicity of AAV vectors significantly limit their application. Here, we endeavored to overcome these limitations by developing a straightforward approach for site-specific PEGylation of AAV via genetic code expansion.

Synthesis of Site-Specific Radiolabeled Antibodies for Radioimmunotherapy via Genetic Code Expansion.

Radioimmunotherapy (RIT) delivers radioisotopes to antigen-expressing cells via monoantibodies for the imaging of lesions or medical therapy. The chelates are typically conjugated to the antibody through cysteine or lysine residues, resulting in heterogeneous chelate-to-antibody ratios and various conjugation sites. To overcome this heterogeneity, we have developed an approach for site-specific radiolabeling of antibodies by combination of genetic code expansion and click chemistry.

Re-exploration of the Codon Context Effect on Amber Codon-Guided Incorporation of Noncanonical Amino Acids in Escherichia coli by the Blue-White Screening Assay.

The effect of codon context on amber codon-guided incorporation of noncanonical amino acids (NAAs) has been previously examined by antibiotic selection. Here, we re-explored this effect by screening a library in which three nucleotides upstream and downstream of the amber codon were randomised, and inserted within the lacZ-α gene. Thousands of clones were obtained and distinguished by the depth of blue colour upon exposure to X-gal.

Data on interaction between adeno-associated virus and U87 cell via cRGD chemical modification.

RGD tripeptide is a specific, high-affinity ligand for integrin, which is highly expressed in cancer cells. We previously reported that cRGD chemically modified AAV2 (AAV2(N587+1/azido+RGD)) showed significantly enhanced infectivity compared to RGD genetically inserted AAV2 (AAV2(N587+RGD)) (10.1016/j.

CRISPRi-Manipulation of Genetic Code Expansion via RF1 for Reassignment of Amber Codon in Bacteria.

The precise engineering of proteins in bacteria via the amber codon has been hampered by the poor incorporation of unnatural amino acid (UAA). Here we explored the amber assignment as a sense codon for UAA by CRISPRi targeting release factor 1 (RF1). Scanning of RF1 gene with sgRNAs identified target loci that differentiate RF1 repressions.

Development of next generation adeno-associated viral vectors capable of selective tropism and efficient gene delivery.

Virus-based nanoparticles have shown promise as vehicles for delivering therapeutic genes. However, the rational design of viral vectors that enable selective tropism towards particular types of cells and tissues remains challenging. Here, we explored structural-functional relationships of the adeno-associated virus 2 (AAV2) vector by expanding its genetic code during production.