A novel host-vector system for direct selection of recombinant baculoviruses (bacmids) in Escherichia coli

Leusch, Mark S.; Lee, Stephen C.; Olins, Peter O.

doi:10.1016/0378-1119(95)00233-v

Cited by 28 publications

(15 citation statements)

References 4 publications

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“…The integration properties of Tn7 have already been used for some biotechnological applications [117,118]. Although it requires four proteins for transposition, the Tn7 system remains interesting for gene therapy purposes because of the occurrence of attTn7 sites downstream eukaryotic genes.…”

Section: Tn7mentioning

confidence: 99%

Sustained Transgene Expression Using Non-Viral Enzymatic Systems for Stable Chromosomal Integration

Palazzoli¹,

Carnus²,

Wells³

et al. 2008

CGT

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Gene delivery technologies have been developed for various biotechnology applications. In gene therapy, they are promising for the treatment of several inherited and acquired human diseases. When therapies require the transfection of a transgene, the vector integration is one of the solutions that is used for maintaining and sustaining expression. On the basis of their origin, vectorisation technologies are currently divided in two fields, gathering on one hand viral vectors and, on the other hand, non-viral approaches. In the case of the non-viral therapies, three main sub-fields are in progress to integrate transgenes. The first uses oligonucleotides to stimulate targeted gene repair by homologous recombination. The second is based on site-specific endonucleases for which the cleavage activity is used to stimulate the host recombination mechanisms in the presence of a DNA vector. The third one is developed from phage and transposon enzymatic systems. The two lasts sub-fields use non-viral enzymes and are the scope of this review. Here, our objective was to overview the main non-viral enzymatic systems able to integrate DNA cassettes. Their molecular and functional characteristics are summarized, and their properties and limits in the current state of the art highlighted. An overview of the safety and quality issues is also presented and discussed, taking into account the solutions that might circumvent problems, intellectual property and economic status for each system. As a conclusion, we propose projections of the future technological developments in the context of the different interests for public and private bodies.

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Section: Tn7mentioning

confidence: 99%

Sustained Transgene Expression Using Non-Viral Enzymatic Systems for Stable Chromosomal Integration

Palazzoli¹,

Carnus²,

Wells³

et al. 2008

CGT

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“…We used retransformation of retrofitted BAC DNA to select for the presence of the Gen marker in the BAC molecule. Alternatively, the library clones could be transferred to a host strain, in which the chromosomal attTn7 site is occupied (Leusch et al, 1995), prior to retrofitting of the BAC clones. The highly specific Tn7-based retrofitting of library clones would permit the addition of genetic elements for the maintenance of the BACs in mammalian cells.…”

Section: Resultsmentioning

confidence: 99%

“…Such transformations, with about 40 ng of the PCR product (not differentiated between linear and circularized molecules), gave rise to about 1000 chloramphenicol-and gentamicin-resistant colonies. Because the Gen gene could be inserted into the attTn7 site of either the BAC molecule or the host chromosome (Leusch et al, 1995), the BAC clones carrying the Gen marker were selected by retransformation into E. coli. Pooled DNA from the initial transformants was used in two parallel transformations, resulting in eight chloramphenicol-and gentamicinresistant clones.…”

Section: Resultsmentioning

confidence: 99%

A Modular, Positive Selection Bacterial Artificial Chromosome Vector with Multiple Cloning Sites

et al. 1999

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“…We have argued the value of a systems engineering perspective on biomedical nanotechnology (Bhalerao et al 2005;), which we illustrated using a molecular nanodevice (bacmid, Leusch et al 1995;Luckow et al 1993). Polyepitope vaccines can be considered multicomponent molecular nanodevices, wherein authentic epitopes are components whose function is immunogenicity and spacers are components whose functions are (minimally) to isolate the authentic epitopes, delimiting immune responses to the authentic epitopes.…”

Section: Discussionmentioning

confidence: 99%

Estimating design space available for polyepitopes through consideration of major histocompatibility complex binding motifs

Lee

Ferrari

Lee

2009

Biomed Microdevices

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Major histocompatibility complex (MHC ) epitope presentation is needed for robust adaptive immune responses. Core peptide binding motifs for class I and class II MHC are 8-10 amino acids long, containing two or more "anchor" residues. These binding motifs define epitope anchor amino acid content and spacing, and knowledge of them has facilitated emergence of polyepitope vaccines. However, polyepitopes can exhibit "junctional epitopes" (neoepitopes interfering with vaccine function) resulting from juxtaposition of authentic epitopes. We have developed an algorithm for consideration of polyepitope sequence in light of MHC motifs to exhaustively identify all junctional-free polyepitope designs for any given set of authentic epitopes, and in so doing discovered that the number of such variants of any given polyepitope can be astronomically high. Our approach designs polyepitopes of any length, considers multiple MHC class I or class II motifs simultaneously and can be adapted to design variants of existing proteins with pre-selected epitope contents. We have also implemented the algorithm as a computer-based tool (CANVAC II), which we make available to interested parties. The vast diversity of junctional-free polyepitopes suggests that the number of potential T-helper epitope free protein variants may also be large, which may have implications for discovery of bioactive but non-immunogenic therapeutics.

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A novel host-vector system for direct selection of recombinant baculoviruses (bacmids) in Escherichia coli

Cited by 28 publications

References 4 publications

Sustained Transgene Expression Using Non-Viral Enzymatic Systems for Stable Chromosomal Integration

Sustained Transgene Expression Using Non-Viral Enzymatic Systems for Stable Chromosomal Integration

A Modular, Positive Selection Bacterial Artificial Chromosome Vector with Multiple Cloning Sites

Estimating design space available for polyepitopes through consideration of major histocompatibility complex binding motifs

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