BioRxiv. 2021 Aug 26
Guochao Liao, Xingxing Fan, Hungyan Lau, Zhongqiu Liu, Chinyu Li, Zeping Xu, Yu Zhang, Xiaoxiao Qi, Dan Li, Qing Zhu, Liqing Chen, Hua Zhou, Sisi Zhu, Bixia Ke, Hudan Pan, Zhe Cong, Yongchao Li, Qian Feng, Qi Lv, Jiangning Liu, Dan Liang, An’an Li, Wenshan Hong, Yebo Li, Linlin Bao, Feng Zhou, Hongbin Gao, Shi Liang, Bihong Huang, Miaoli Wu, Chuan Qin, Changwen Ke, Liang Liu
Products used in the paper Details Operation
plasmid construction & AAV vector packaging The recombinant type 5 adenoviral-vectored SARS-CoV-2 vaccines encoding RBD domain (residues 319-541), RBD-plus domain (residues 319-583), S1 protein, full-length of S protein and NTD domain of SARS-CoV-2 were produced by PackGene Biotech (Guangzhou, China). Briefly, rAAV5 packaging plasmids were transfected into HEK293T cells using PEI transfection reagent, according to the manufacturer’s protocol. The transfected cells and supernatants were harvested 72 h post transfection. rAAV5-vaccine was purified and titrated by real-time quantitative PCR (Q- PCR). rAAV5-vaccine was adjusted to 1012 GCs/ml in PBS and used for the following vaccinations. Request Quote

Research Field: Covid-related

AAV Serotype: AAV5

Dose: All animals were immunized with rAAV5-based vaccines or rAAV5-GFP via i.m in the hind leg or via intranasal inoculation at a single dose (five to eight animals for each group).

Animal or cell line strain: 6 weeks-old, 36 weeks-old female BALB/c mice and 6-8 weeks female Wistar rats were purchased from the Laboratory Animal Center of Southern Medical University (Guangdong, China).


The COVID-19 pandemic and the SARS-CoV-2 with its variants have posed unprecedented challenges worldwide. Existing vaccines have limited effectiveness against the SARS-CoV-2 variants. Therefore, novel vaccines to match current mutated viral lineages with long-term protective immunity are urgently in demand. In the current study, we for the first time designed a recombinant Adeno-Associated Virus 5 (rAAV5)-based vaccine named as rAAV-COVID-19 vaccine (Covacinplus) by using RBD-plus of spike protein with both the single-stranded and the self-complementary AAV5 delivering vectors (ssAAV5 and scAAAV5), which provides excellent protection from SARS-CoV-2 infection. A single dose vaccination induced the strong immune response against SARS-CoV-2. The induced neutralizing antibodies (NAs) titers were maintained at a high peak level of over 1:1024 even after more than one year of injection and accompanied with functional T-cells responses in mice. Importantly, both ssAAV- and scAAV-based RBD-plus vaccines exhibited high levels of serum NAs against current circulating variants including variants Alpha, Beta, Gamma and Delta. SARS-CoV-2 virus challenge test showed that ssAAV5-RBD-plus vaccine protected both young and old age mice from SARS-CoV-2 infection in the upper and the lower respiratory tracts. Moreover, whole genome sequencing demonstrated that AAV vector DNA sequences were not found in the genome of the vaccinated mice after one year vaccination, demonstrating excellent safety of the vaccine. Taken together, this study suggests that rAAV5-based vaccine is powerful against SARS-CoV-2 and its variants with long-term protective immunity and excellent safety, which has great potential for development into prophylactic vaccination in human to end this global pandemic.

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