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2.
Vaccine ; 40(49): 7022-7031, 2022 Nov 22.
Article in English | MEDLINE | ID: covidwho-2119102

ABSTRACT

Historically, virulent variola virus infection caused hundreds of millions of deaths. The smallpox pandemic in human beings has spread for centuries until the advent of the attenuated vaccinia virus (VV) vaccine, which played a crucial role in eradicating the deadly contagious disease. Decades of exploration and utilization have validated the attenuated VV as a promising vaccine vehicle against various lethal viruses. In this review, we focus on the advances in VV-based vaccine vector studies, including construction approaches of recombinant VV, the impact of VV-specific pre-existing immunity on subsequent VV-based vaccines, and antigen-specific immune responses. More specifically, the recombinant VV-based flaviviruses are intensively discussed. Based on the publication data, this review aims to provide valuable insights and guidance for future VV-based vaccine development.


Subject(s)
Flavivirus , Smallpox Vaccine , Vaccines , Vaccinia , Humans , Vaccinia virus , Flavivirus/genetics , Vaccine Development , Genetic Vectors
3.
Nat Biotechnol ; 40(11): 1586-1600, 2022 Nov.
Article in English | MEDLINE | ID: covidwho-2106427

ABSTRACT

The extraordinary success of mRNA vaccines against coronavirus disease 2019 (COVID-19) has renewed interest in mRNA as a means of delivering therapeutic proteins. Early clinical trials of mRNA therapeutics include studies of paracrine vascular endothelial growth factor (VEGF) mRNA for heart failure and of CRISPR-Cas9 mRNA for a congenital liver-specific storage disease. However, a series of challenges remains to be addressed before mRNA can be established as a general therapeutic modality with broad relevance to both rare and common diseases. An array of new technologies is being developed to surmount these challenges, including approaches to optimize mRNA cargos, lipid carriers with inherent tissue tropism and in vivo percutaneous delivery systems. The judicious integration of these advances may unlock the promise of biologically targeted mRNA therapeutics, beyond vaccines and other immunostimulatory agents, for the treatment of diverse clinical indications.


Subject(s)
Genetic Vectors , RNA, Messenger , Humans , COVID-19/prevention & control , RNA, Messenger/genetics , RNA, Messenger/therapeutic use , Vascular Endothelial Growth Factor A/genetics , Vascular Endothelial Growth Factor A/metabolism , COVID-19 Vaccines
4.
Genes (Basel) ; 13(11)2022 10 25.
Article in English | MEDLINE | ID: covidwho-2090054

ABSTRACT

Adenoviral vaccines have been at the front line in the fight against pandemics caused by viral infections such as Ebola and the coronavirus disease 2019. This has revived an interest in developing these vectors as vaccines and therapies against other viruses of health importance such as hepatitis B virus (HBV). Current hepatitis B therapies are not curative; hence, chronic hepatitis B remains the major risk factor for development of liver disease and death in HBV-infected individuals. The ability to induce a robust immune response and high liver transduction efficiency makes adenoviral vectors attractive tools for anti-HBV vaccine and therapy development, respectively. This review describes recent developments in designing adenoviral-vector-based therapeutics and vaccines against HBV infection.


Subject(s)
COVID-19 , Hepatitis B, Chronic , Hepatitis B , Viral Vaccines , Humans , Genetic Vectors/genetics , Hepatitis B virus/genetics , Hepatitis B/genetics , Hepatitis B/prevention & control
5.
Viruses ; 14(10)2022 09 30.
Article in English | MEDLINE | ID: covidwho-2066551

ABSTRACT

Molecular therapies exploiting mRNA vectors embody enormous potential, as evidenced by the utility of this technology for the context of the COVID-19 pandemic. Nonetheless, broad implementation of these promising strategies has been restricted by the limited repertoires of delivery vehicles capable of mRNA transport. On this basis, we explored a strategy based on exploiting the well characterized entry biology of adenovirus. To this end, we studied an adenovirus-polylysine (AdpL) that embodied "piggyback" transport of the mRNA on the capsid exterior of adenovirus. We hypothesized that the efficient steps of Ad binding, receptor-mediated entry, and capsid-mediated endosome escape could provide an effective pathway for transport of mRNA to the cellular cytosol for transgene expression. Our studies confirmed that AdpL could mediate effective gene transfer of mRNA vectors in vitro and in vivo. Facets of this method may offer key utilities to actualize the promise of mRNA-based therapeutics.


Subject(s)
Adenoviridae Infections , COVID-19 , Humans , Adenoviridae/genetics , Genetic Vectors/genetics , Gene Transfer Techniques , Polylysine , RNA, Messenger/genetics , RNA, Messenger/metabolism , Pandemics , Capsid Proteins/genetics , Capsid Proteins/metabolism , Biology
6.
Viruses ; 14(9)2022 09 02.
Article in English | MEDLINE | ID: covidwho-2055390

ABSTRACT

Only two decades after discovering miRNAs, our understanding of the functional effects of deregulated miRNAs in the development of diseases, particularly cancer, has been rapidly evolving. These observations and functional studies provide the basis for developing miRNA-based diagnostic markers or new therapeutic strategies. Adenoviral (Ad) vectors belong to the most frequently used vector types in gene therapy and are suitable for strong short-term transgene expression in a variety of cells. Here, we report the set-up and functionality of an Ad-based miRNA vector platform that can be employed to deliver and express a high level of miRNAs efficiently. This vector platform allows fast and efficient vector production to high titers and the expression of pri-miRNA precursors under the control of a polymerase II promoter. In contrast to non-viral miRNA delivery systems, this Ad-based miRNA vector platform allows accurate dosing of the delivered miRNAs. Using a two-vector model, we showed that Ad-driven miRNA expression was sufficient in down-regulating the expression of an overexpressed and highly stable protein. Additional data corroborated the downregulation of multiple endogenous target RNAs using the system presented here. Additionally, we report some unanticipated synergistic effects on the transduction efficiencies in vitro when cells were consecutively transduced with two different Ad-vectors. This effect might be taken into consideration for protocols using two or more different Ad vectors simultaneously.


Subject(s)
MicroRNAs , Adenoviridae/genetics , Adenoviridae/metabolism , Genetic Therapy/methods , Genetic Vectors/genetics , MicroRNAs/genetics , MicroRNAs/metabolism , Transgenes
7.
Curr Opin HIV AIDS ; 17(6): 338-344, 2022 11 01.
Article in English | MEDLINE | ID: covidwho-2051602

ABSTRACT

PURPOSE OF REVIEW: The purpose of this review is to share the excitement of new developments in the field of vaccine vector modalities against infectious diseases. The focus is on HIV-1/AIDS with reference to the most successful as well as currently tested COVID-19 vaccines, and human trials, which best inform iterative vaccine improvements. RECENT FINDINGS: Several genetic subunit vaccines against SARS-CoV-2 demonstrated protection against severe disease, obtained Emergency Use Authorization and scaled their production to billions of doses. Many more are in efficacy evaluation. In contrast, development of HIV-1 vaccines has been extremely difficult. Perseverance of scientists is deepening our understanding of what constitutes immunity against HIV-1 infection and how to achieve protective levels of relevant responses by active immunization, passive administration or a combination of both. Novel platforms led by RNA play a pivotal role. However, a difficult virus may require a complex approach. Proof of concept for HIV-1 prevention and cure might be at reach, and when it arrives, it will be a great and needed encouragement to the field. SUMMARY: Despite the enormous success of drug treatment, vaccines remain the best solution and likely a necessary component of any package that truly ends the AIDS epidemic.


Subject(s)
AIDS Vaccines , Acquired Immunodeficiency Syndrome , COVID-19 , HIV Infections , Viral Vaccines , AIDS Vaccines/genetics , COVID-19/prevention & control , COVID-19 Vaccines/genetics , DNA, Viral , Genetic Vectors , HIV Infections/prevention & control , Humans , RNA, Messenger , SARS-CoV-2/genetics , Vaccines, Subunit , Viral Vaccines/genetics
8.
Mol Neurobiol ; 59(1): 191-233, 2022 Jan.
Article in English | MEDLINE | ID: covidwho-2041322

ABSTRACT

The etiology of many neurological diseases affecting the central nervous system (CNS) is unknown and still needs more effective and specific therapeutic approaches. Gene therapy has a promising future in treating neurodegenerative disorders by correcting the genetic defects or by therapeutic protein delivery and is now an attraction for neurologists to treat brain disorders, like Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, spinal muscular atrophy, spinocerebellar ataxia, epilepsy, Huntington's disease, stroke, and spinal cord injury. Gene therapy allows the transgene induction, with a unique expression in cells' substrate. This article mainly focuses on the delivering modes of genetic materials in the CNS, which includes viral and non-viral vectors and their application in gene therapy. Despite the many clinical trials conducted so far, data have shown disappointing outcomes. The efforts done to improve outcomes, efficacy, and safety in the identification of targets in various neurological disorders are also discussed here. Adapting gene therapy as a new therapeutic approach for treating neurological disorders seems to be promising, with early detection and delivery of therapy before the neuron is lost, helping a lot the development of new therapeutic options to translate to the clinic.


Subject(s)
Genetic Therapy , Intercellular Signaling Peptides and Proteins/genetics , Neurodegenerative Diseases/therapy , Genetic Vectors , Humans , Neurodegenerative Diseases/genetics , Treatment Outcome
9.
Am J Ophthalmol ; 241: 262-271, 2022 09.
Article in English | MEDLINE | ID: covidwho-2014767

ABSTRACT

PURPOSE: To assess safety of gene therapy in G11778A Leber hereditary optic neuropathy (LHON). DESIGN: Phase 1 clinical trial. METHODS: Setting: single institution. PARTICIPANTS: Patients with G11778A LHON and chronic bilateral visual loss >12 months (group 1, n = 11), acute bilateral visual loss <12 months (group 2, n = 9), or unilateral visual loss (group 3, n = 8). INTERVENTION: unilateral intravitreal AAV2(Y444,500,730F)-P1ND4v2 injection with low, medium, high, and higher doses to worse eye for groups 1 and 2 and better eye for group 3. OUTCOME MEASURES: Best-corrected visual acuity (BCVA), adverse events, and vector antibody responses. Mean follow-up was 24 months (range, 12-36 months); BCVAs were compared with a published prospective natural history cohort with designated surrogate study and fellow eyes. RESULTS: Incident uveitis (8 of 28, 29%), the only vector-related adverse event, resulted in no attributable vision sequelae and was related to vector dose: 5 of 7 (71%) higher-dose eyes vs 3 of 21 (14%) low-, medium-, or high-dose eyes (P < .001). Incident uveitis requiring treatment was associated with increased serum AAV2 neutralizing antibody titers (p=0.007) but not serum AAV2 polymerase chain reaction. Improvements of ≥15-letter BCVA occurred in some treated and fellow eyes of groups 1 and 2 and some surrogate study and fellow eyes of natural history subjects. All study eyes (BCVA ≥20/40) in group 3 lost ≥15 letters within the first year despite treatment. CONCLUSIONS: G11778A LHON gene therapy has a favorable safety profile. Our results suggest that if there is an efficacy effect, it is likely small and not dose related. Demonstration of efficacy requires randomization of patients to a group not receiving vector in either eye.


Subject(s)
Optic Atrophy, Hereditary, Leber , DNA, Mitochondrial/genetics , Dependovirus/genetics , Dependovirus/metabolism , Electroretinography , Genetic Therapy/adverse effects , Genetic Therapy/methods , Genetic Vectors , Humans , NADH Dehydrogenase/genetics , NADH Dehydrogenase/metabolism , Optic Atrophy, Hereditary, Leber/genetics , Optic Atrophy, Hereditary, Leber/therapy , Prospective Studies , Retinal Ganglion Cells , Tomography, Optical Coherence , Vision Disorders/etiology , Visual Acuity , Visual Fields
10.
Viruses ; 12(1)2020 01 20.
Article in English | MEDLINE | ID: covidwho-1969491

ABSTRACT

Middle East respiratory syndrome (MERS) is an acute, high-mortality-rate, severe infectious disease caused by an emerging MERS coronavirus (MERS-CoV) that causes severe respiratory diseases. The continuous spread and great pandemic potential of MERS-CoV make it necessarily important to develop effective vaccines. We previously demonstrated that the application of Gram-positive enhancer matrix (GEM) particles as a bacterial vector displaying the MERS-CoV receptor-binding domain (RBD) is a very promising MERS vaccine candidate that is capable of producing potential neutralization antibodies. We have also used the rabies virus (RV) as a viral vector to design a recombinant vaccine by expressing the MERS-CoV S1 (spike) protein on the surface of the RV. In this study, we compared the immunological efficacy of the vaccine candidates in BALB/c mice in terms of the levels of humoral and cellular immune responses. The results show that the rabies virus vector-based vaccine can induce remarkably earlier antibody response and higher levels of cellular immunity than the GEM particles vector. However, the GEM particles vector-based vaccine candidate can induce remarkably higher antibody response, even at a very low dose of 1 µg. These results indicate that vaccines constructed using different vaccine vector platforms for the same pathogen have different rates and trends in humoral and cellular immune responses in the same animal model. This discovery not only provides more alternative vaccine development platforms for MERS-CoV vaccine development, but also provides a theoretical basis for our future selection of vaccine vector platforms for other specific pathogens.


Subject(s)
Coronavirus Infections/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , Viral Vaccines/immunology , Animals , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Cell Line , Coronavirus Infections/prevention & control , Genetic Vectors , Humans , Immunization , Immunoglobulin G/blood , Immunoglobulin G/immunology , Lactococcus lactis/genetics , Mice , Mice, Inbred BALB C , Middle East Respiratory Syndrome Coronavirus/genetics , Rabies virus/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/immunology , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/immunology , Viral Vaccines/administration & dosage
12.
Int J Mol Sci ; 23(14)2022 Jul 07.
Article in English | MEDLINE | ID: covidwho-1928572

ABSTRACT

Sleeping Beauty (SB) is the first DNA transposon employed for efficient transposition in vertebrate cells, opening new applications for genetic engineering and gene therapies. A transposon-based gene delivery system holds the favourable features of non-viral vectors and an attractive safety profile. Here, we employed SB to engineer HEK293 cells for optimizing the production of a chimpanzee Adenovector (chAd) belonging to the Human Mastadenovirus C species. To date, chAd vectors are employed in several clinical settings for infectious diseases, last but not least COVID-19. A robust, efficient and quick viral vector production could advance the clinical application of chAd vectors. To this aim, we firstly swapped the hAd5 E1 with chAd-C E1 gene by using the CRISPR/Cas9 system. We demonstrated that in the absence of human Ad5 E1, chimp Ad-C E1 gene did not support HEK293 survival. To improve chAd-C vector production, we engineered HEK293 cells to stably express the chAd-C precursor terminal protein (ch.pTP), which plays a crucial role in chimpanzee Adenoviral DNA replication. The results indicate that exogenous ch.pTP expression significantly ameliorate the packaging and amplification of recombinant chAd-C vectors thus, the engineered HEK293ch.pTP cells could represent a superior packaging cell line for the production of these vectors.


Subject(s)
COVID-19 , Pan troglodytes , Adenoviridae/genetics , Animals , DNA Transposable Elements/genetics , Gene Transfer Techniques , Genetic Vectors/genetics , HEK293 Cells , Humans , Pan troglodytes/genetics
13.
ACS Synth Biol ; 11(7): 2229-2237, 2022 07 15.
Article in English | MEDLINE | ID: covidwho-1921556

ABSTRACT

Rapid and flexible plasmid construct generation at scale is one of the most limiting first steps in drug discovery projects. These hurdles can partly be overcome by adopting modular DNA design principles, automated sequence fragmentation, and plasmid assembly. To this end we have designed a robust, multimodule golden gate based cloning platform for construct generation with a wide range of applications. The assembly efficiency of the system was validated by splitting sfGFP and sfCherry3C cassettes and expressing them in E. coli followed by fluorometric assessment. To minimize timelines and cost for complex constructs, we developed a software tool named FRAGLER (FRAGment recycLER) that performs codon optimization, multiple sequence alignment, and automated generation of fragments for recycling. To highlight the flexibility and robustness of the platform, we (i) generated plasmids for SarsCoV2 protein reagents, (ii) automated and parallelized assemblies, and (iii) built modular libraries of chimeric antigen receptors (CARs) variants. Applying the new assembly framework, we have greatly streamlined plasmid construction and increased our capacity for rapid generation of complex plasmids.


Subject(s)
COVID-19 , Escherichia coli , Cloning, Molecular , DNA/genetics , Escherichia coli/genetics , Genetic Vectors , Humans , Plasmids/genetics , RNA, Viral , SARS-CoV-2 , Synthetic Biology
14.
Curr Opin Immunol ; 77: 102210, 2022 08.
Article in English | MEDLINE | ID: covidwho-1920777

ABSTRACT

Over the past two years, the SARS-CoV-2 pandemic has highlighted the impact that emerging pathogens can have on global health. The development of new and effective vaccine technologies is vital in the fight against such threats. Viral vectors are a relatively new vaccine platform that relies on recombinant viruses to deliver selected immunogens into the host. In response to the SARS-CoV-2 pandemic, the development and subsequent rollout of adenoviral vector vaccines has shown the utility, impact, scalability and efficacy of this platform. Shown to elicit strong cellular and humoral immune responses in diverse populations, these vaccine vectors will be an important approach against infectious diseases in the future.


Subject(s)
COVID-19 , Viral Vaccines , COVID-19/prevention & control , Genetic Vectors/genetics , Humans , Pandemics/prevention & control , SARS-CoV-2
15.
J Infect Dis ; 226(6): 979-982, 2022 09 21.
Article in English | MEDLINE | ID: covidwho-1886441

ABSTRACT

This secondary analysis of the phase 3 ENSEMBLE trial (NCT04505722) assessed the impact of preexisting humoral immunity to adenovirus 26 (Ad26) on the immunogenicity of Ad26.COV2.S-elicited severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibody levels in 380 participants in Brazil, South Africa, and the United States. Among those vaccinated in Brazil and South Africa, 31% and 66%, respectively, had prevaccination serum-neutralizing activity against Ad26, with little preexisting immunity detected in the United States. Vaccine recipients in each country had similar postvaccination spike (S) protein-binding antibody levels, indicating that baseline immunity to Ad26 has no clear impact on vaccine-induced immune responses.


Subject(s)
Adenoviridae Infections , COVID-19 , Ad26COVS1 , Adenoviridae , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/prevention & control , COVID-19 Vaccines , Genetic Vectors , Humans , Immunity, Cellular , Immunity, Humoral , Immunogenicity, Vaccine , SARS-CoV-2
17.
J Gen Virol ; 103(5)2022 05.
Article in English | MEDLINE | ID: covidwho-1861028

ABSTRACT

In vivo nucleic expression technologies using DNA or mRNA offer several advantages for recombinant gene expression. Their inherent ability to generate natively expressed recombinant proteins and antigens allows these technologies to mimic foreign gene expression without infection. Furthermore, foreign nucleic acid fragments have an inherent ability to act as natural immune adjuvants and stimulate innate pathogen- and DNA damage-associated receptors that are responsible for activating pathogen-associated molecular pattern (PAMP) and DNA damage-associated molecular pattern (DAMP) signalling pathways. This makes nucleic-acid-based expression technologies attractive for a wide range of vaccine and oncolytic immunotherapeutic uses. Recently, RNA vaccines have demonstrated their efficacy in generating strong humoral and cellular immune responses for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). DNA vaccines, which are more stable and easier to manufacture, generate similar immune responses to RNA, but typically exhibit lower immunogenicity. Here we report on a novel method of constructing self-amplifying DNA expression vectors that have the potential to amplify and enhance gene/antigen expression at a cellular level by increasing per cell gene copy numbers, boost genomic adjuvating effects and mitigate through replication many of the problems faced by non-replicating vectors such as degradation, methylation and gene silencing. These vectors employ a viral origin rolling circle replication cycle in mammalian host cells that amplifies the vector and gene of interest (GOI) copy number, maintaining themselves as nuclear episomes. We show that these vectors maintain persistently elevated GOI expression levels at the cellular level and induce morphological cellular alterations synonymous with increased cellular stress.


Subject(s)
COVID-19 , Circovirus , Vaccines, DNA , Animals , Circovirus/genetics , Genetic Vectors/genetics , Mammals , SARS-CoV-2 , Vaccines, DNA/genetics
18.
J Mol Med (Berl) ; 100(6): 875-901, 2022 06.
Article in English | MEDLINE | ID: covidwho-1858961

ABSTRACT

Despite the development of novel pharmacological treatments, cardiovascular disease morbidity and mortality remain high indicating an unmet clinical need. Viral gene therapy enables targeted delivery of therapeutic transgenes and represents an attractive platform for tackling acquired and inherited cardiovascular diseases in the future. Current cardiovascular gene therapy trials in humans mainly focus on improving cardiac angiogenesis and function. Encouragingly, local delivery of therapeutic transgenes utilising first-generation human adenovirus serotype (HAd)-5 is safe in the short term and has shown some efficacy in drug refractory angina pectoris and heart failure with reduced ejection fraction. Despite this success, systemic delivery of therapeutic HAd-5 vectors targeting cardiovascular tissues and internal organs is limited by negligible gene transfer to target cells, elimination by the immune system, liver sequestration, off-target effects, and episomal degradation. To circumvent these barriers, cardiovascular gene therapy research has focused on determining the safety and efficacy of rare alternative serotypes and/or genetically engineered adenoviral capsid protein-modified vectors following local or systemic delivery. Pre-clinical studies have identified several vectors including HAd-11, HAd-35, and HAd-20-42-42 as promising platforms for local and systemic targeting of vascular endothelial and smooth muscle cells. In the past, clinical gene therapy trials were often restricted by limited scale-up capabilities of gene therapy medicinal products (GTMPs) and lack of regulatory guidance. However, significant improvement of industrial GTMP scale-up and purification, development of novel producer cell lines, and issuing of GTMP regulatory guidance by national regulatory health agencies have addressed many of these challenges, creating a more robust framework for future adenoviral-based cardiovascular gene therapy. In addition, this has enabled the mass roll out of adenovirus vector-based COVID-19 vaccines. KEY MESSAGES: First-generation HAd-5 vectors are widely used in cardiovascular gene therapy. HAd-5-based gene therapy was shown to lead to cardiac angiogenesis and improved function. Novel HAd vectors may represent promising transgene carriers for systemic delivery. Novel methods allow industrial scale-up of rare/genetically altered Ad serotypes. National regulatory health agencies have issued guidance on GMP for GTMPs.


Subject(s)
COVID-19 , Gene Transfer Techniques , Adenoviridae/genetics , COVID-19 Vaccines , Genetic Therapy/methods , Genetic Vectors/genetics , Humans
20.
Viruses ; 14(3)2022 03 06.
Article in English | MEDLINE | ID: covidwho-1765949

ABSTRACT

Gene therapy and vaccine development need more novel adenovirus vectors. Here, we attempt to provide strategies to construct adenovirus vectors based on restriction-assembly for researchers with little experience in this field. Restriction-assembly is a combined method of restriction digestion and Gibson assembly, by which the major part of the obtained plasmid comes from digested DNA fragments instead of PCR products. We demonstrated the capability of restriction-assembly in manipulating the genome of simian adenovirus 1 (SAdV-1) in this study. A PCR product of the plasmid backbone was combined with SAdV-1 genomic DNA to construct an infectious clone, plasmid pKSAV1, by Gibson assembly. Restriction-assembly was performed repeatedly in the steps of intermediate plasmid isolation, modification, and restoration. The generated adenoviral plasmid was linearized by restriction enzyme digestion and transfected into packaging 293 cells to rescue E3-deleted replication-competent SAdV1XE3-CGA virus. Interestingly, SAdV1XE3-CGA could propagate in human chronic myelogenous leukemia K562 cells. The E1 region was similarly modified to generate E1/E3-deleted replication-defective virus SAdV1-EG. SAdV1-EG had a moderate gene transfer ability to adherent mammalian cells, and it could efficiently transduce suspension cells when compared with the human adenovirus 5 control vector. Restriction-assembly is easy to use and can be performed without special experimental materials and instruments. It is highly effective with verifiable outcomes at each step. More importantly, restriction-assembly makes the established vector system modifiable, upgradable and under sustainable development, and it can serve as the instructive method or strategy for the synthetic biology of adenoviruses.


Subject(s)
Adenoviruses, Human , Adenoviruses, Simian , Adenoviridae/genetics , Adenoviruses, Human/genetics , Adenoviruses, Simian/genetics , Animals , DNA , Genetic Vectors/genetics , Humans , Mammals
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