Adenoviral Vector Development Service
Recombinant adenovirus vectors have been widely used to deliver exogenous genes to a variety of cell types and tissues both in vitro and in vivo. They can be easily grown to high titer and can efficiently transfer genes into both dividing and non-dividing cells. Over the years, numerous approaches have been developed for the construction of recombinant adenoviruses. As a leading company in this field, Creative Biolabs provides the state-of-the-art recombinant adenoviral vector construction service to meet the demands in basic research and preclinical applications.
Recombinant Adenovirus Rescue in Mammalian Cells
Mammalian cells, such as 293 cells, have long been used to generate recombinant adenovirus vectors. This method uses two plasmids coding for the homologous recombinant regions: one is a shuttle plasmid containing an expression cassette, and the other is the large plasmid containing the majority of the adenoviral genome. After the gene of interest has been introduced into the second plasmid, the two plasmids are co-transfected into mammalian cells, and the virus produced by recombination in 293 cells is isolated through plaque purification.
Figure 1. Schematic outline of the AdEasy system. (He, 1998)
Recombinant Adenovirus Construction in Bacterial Systems
In bacterial systems, no ligation steps are needed in generating the plasmids, and any region of the adenoviral genome can be easily modified. Briefly, the full-length adenoviral genome flanked by unique restriction enzyme sites is firstly cloned into a bacterial plasmid. Next, the region of the viral genome to be modified is subcloned into a bacterial shuttle plasmid, and the desired changes are introduced by molecular biology techniques.
Capsid-modified Adenovirus Vector Construction
Adenovirus infection is dependent on the expression levels of the coxsackievirus and adenovirus receptor (CAR) in target cells which means that Ad vectors cannot transfer genes of interest into cells lacking CAR expression (i.e. many advanced tumor cells, peripheral blood cells, hematopoietic stem cells, dendritic cells, etc.). To overcome this difficulty, capsid-modified Ad vector which is constructed by genetic modification of the Ad capsid, such as its fiber, protein IX (pIX), or hexon, has been developed as an attractive strategy for altering the Ad tropism.
Regulated Adenovirus Vector Construction
Control of gene expression using tissue- and cell-specific promoters have been tested extensively as a means of targeting transgene expression. Here, we are able to construct a regulatable adenovirus vector to help artificially control gene expression both in vitro and in vivo.
Recombinant Adenovirus Vector Design for Invading Immune System
As viruses have evolved to more efficiently transduce host cells, the mammalian immune system has coevolved in cellular and humoral immune responses to prevent the growth of an invading virus or pathogen. Particularly, activation of the innate and adaptive immune systems by adenovirus vectors can lead to tissue or organ inflammation, enhanced immune-mediated clearance of vector transduced cells, and reduced transgene expression.
Pseudotyping Adenoviral Vectors Construction
While adenovirus gene delivery vectors are useful in many gene therapy applications, their broad tropism means that they cannot be directed to a specific target cell. However, pseudotyped adenoviral vectors that are not prone to neutralization have high in vivo target specificity.
Adenoviral/Retroviral Hybrid Vector Construction
The hybrid adenoviral/retroviral vector system is designed to incorporate favorable aspects of both adenoviral and retroviral vectors. In this system, the non-integrative adenoviral vector is rendered functionally integrative via the intermediate generation of a retroviral producer cell. The chimeric vectors may allow the realization of the requisite goals for specific gene-therapy applications.
Ad/AAV Hybrid Vectors Construction
Adenovirus (Ad) and adeno-associated virus (AAV) have attractive and complementary properties that can be exploited for gene transfer purposes. The hybrid Ad/AAV vectors are constructed based on coupling the AAV DNA replication mechanism to the Ad encapsidation process through the packaging of AAV-dependent replicative intermediates provided with Ad packaging elements into Ad capsids.
Based on our top technology platform, Creative Biolabs is dedicated to offering comprehensive recombinant adenovirus vector construction services for gene therapy. For more information, please feel free to contact us.
Reference
- He, T.C.; et al. (1998). A simplified system for generating recombinant adenoviruses. Proceedings of the National Academy of Sciences. 95(5): pp.2509-2514.
