scholarly journals DNA-mediated gene transfer without carrier DNA.

1981 ◽  
Vol 91 (1) ◽  
pp. 153-156 ◽  
Author(s):  
K M Huttner ◽  
J A Barbosa ◽  
G A Scangos ◽  
D D Pratcheva ◽  
F H Ruddle
Keyword(s):  
Gene Transfer ◽  
Dna Sequences ◽  
Kinase Gene ◽  
Whole Cell ◽  
Cell Carrier ◽  
Wide Range ◽  
Dna Restriction ◽  
Simplex Virus

DNA-mediated gene transfer is a procedure which uses purified DNA to introduce new genetic elements into cells in culture. The standard DNA-mediated gene transfer procedure involves the use of whole cell DNA as carrier DNA for the transfer. We have modified the standard DNA-mediated gene transfer procedure to transfer the Herpes simplex virus type 1 thymidine kinase gene (TK) into TK- murine recipient cells in the absence of whole cell carrier DNA. The majority (8/10) of carrier-free transformant lines expressed the TK+ phenotype stably, in sharp contrast to our results with carrier-containing DNA-mediated gene transfer. There was a wide range in donor DNA content among independent transformants. Further analysis on one transformant line using DNA restriction digests and in situ hybridization provided evidence that, in the absence of whole cell carrier DNA, multiple donor DNA sequences became integrated at a single chromosomal site.

scholarly journals Adaptation of a retrovirus as a eucaryotic vector transmitting the herpes simplex virus thymidine kinase gene.

1982 ◽  
Vol 2 (4) ◽  
pp. 426-436 ◽  
Author(s):  
C J Tabin ◽  
J W Hoffmann ◽  
S P Goff ◽  
R A Weinberg
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Thymidine Kinase ◽  
Dna Sequences ◽  
Leukemia Virus ◽  
Chimeric Virus ◽  
Animal Cells ◽  
Kinase Gene ◽  
General Applicability ◽  
Simplex Virus

We investigated the feasibility of using retroviruses as vectors for transferring DNA sequences into animal cells. The thymidine kinase (tk) gene of herpes simplex virus was chosen as a convenient model. The internal BamHI fragments of a DNA clone of Moloney leukemia virus (MLV) were replaced with a purified BamHI DNA segment containing the tk gene. Chimeric genomes were created carrying the tk insert in both orientations relative to the MLV sequence. Each was transfected into TK- cells along with MLV helper virus, and TK+ colonies were obtained by selection in the presence of hypoxanthine, aminopterin, and thymidine (HAT). Virus collected from TK+-transformed, MLV producer cells passed the TK+ phenotype to TK- cells. Nonproducer cells were isolated, and TK+ transducing virus was subsequently rescued from them. The chimeric virus showed single-hit kinetics in infections. Virion and cellular RNA and cellular DNA from infected cells were all shown to contain sequences which hybridized to both MLV- and tk-specific probes. The sizes of these sequences were consistent with those predicted for the chimeric virus. In all respects studied, the chimeric MLV-tk virus behaved like known replication-defective retroviruses. These experiments suggest great general applicability of retroviruses as eucaryotic vectors.

Adenoviral-Mediated Herpes Simplex Virus Thymidine Kinase Gene Transfer

Pancreas ◽  
1997 ◽  
Vol 15 (1) ◽  
pp. 25-34 ◽  
Author(s):  
Andreas Block ◽  
Shu-Hsia Chen ◽  
Ken-Ichiro Kosai ◽  
Milton Finegold ◽  
Savio L. C. Woo
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Gene Transfer ◽  
Thymidine Kinase ◽  
Thymidine Kinase Gene ◽  
Kinase Gene ◽  
Simplex Virus

Dominant positive and negative selection using a hygromycin phosphotransferase-thymidine kinase fusion gene

1991 ◽  
Vol 11 (6) ◽  
pp. 3374-3378 ◽  
Author(s):  
S D Lupton ◽  
L L Brunton ◽  
V A Kalberg ◽  
R W Overell
Keyword(s):  
Thymidine Kinase ◽  
Negative Selection ◽  
Fusion Gene ◽  
Hygromycin B ◽  
Thymidine Kinase Gene ◽  
Kinase Gene ◽  
Hygromycin Phosphotransferase ◽  
Simplex Virus ◽  
Genetic Entity

The hygromycin phosphotransferase gene was fused in-frame with the herpes simplex virus type 1 thymidine kinase gene. The resulting fusion gene (termed HyTK) confers hygromycin B resistance for dominant positive selection and ganciclovir sensitivity for negative selection and provides a means by which these selectable phenotypes may be expressed and regulated as a single genetic entity.

scholarly journals Adenoviral-Mediated Herpes Simplex Virus-Thymidine Kinase Gene Transfer in Vivo for Treatment of Experimental Human Melanoma

1996 ◽  
Vol 106 (6) ◽  
pp. 1163-1168 ◽  
Author(s):  
Bernd Bonnekoh ◽  
David A. Greenhalgh ◽  
Donnie S. Bundman ◽  
Ken-ichiro Kosai ◽  
Shu-Hsia Chen ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Gene Transfer ◽  
Thymidine Kinase ◽  
Human Melanoma ◽  
Thymidine Kinase Gene ◽  
Kinase Gene ◽  
Simplex Virus

scholarly journals The 3-O-sulfation of heparan sulfate modulates protein binding and lyase degradation

2021 ◽  
Vol 118 (3) ◽  
pp. e2012935118
Author(s):  
Pradeep Chopra ◽  
Apoorva Joshi ◽  
Jiandong Wu ◽  
Weigang Lu ◽  
Tejabhiram Yadavalli ◽  
...  
Keyword(s):  
Heparan Sulfate ◽  
Binding Proteins ◽  
Factor Xa ◽  
Tissue Expression ◽  
Cell Entry ◽  
Synthetic Approach ◽  
Array Technology ◽  
Wide Range ◽  
Simplex Virus

Humans express seven heparan sulfate (HS) 3-O-sulfotransferases that differ in substrate specificity and tissue expression. Although genetic studies have indicated that 3-O-sulfated HS modulates many biological processes, ligand requirements for proteins engaging with HS modified by 3-O-sulfate (3-OS) have been difficult to determine. In particular, the context in which the 3-OS group needs to be presented for binding is largely unknown. We describe herein a modular synthetic approach that can provide structurally diverse HS oligosaccharides with and without 3-OS. The methodology was employed to prepare 27 hexasaccharides that were printed as a glycan microarray to examine ligand requirements of a wide range of HS-binding proteins. The binding selectivity of antithrombin-III (AT-III) compared well with anti-Factor Xa activity supporting robustness of the array technology. Many of the other examined HS-binding proteins required an IdoA2S-GlcNS3S6S sequon for binding but exhibited variable dependence for the 2-OS and 6-OS moieties, and a GlcA or IdoA2S residue neighboring the central GlcNS3S. The HS oligosaccharides were also examined as inhibitors of cell entry by herpes simplex virus type 1, which, surprisingly, showed a lack of dependence of 3-OS, indicating that, instead of glycoprotein D (gD), they competitively bind to gB and gC. The compounds were also used to examine substrate specificities of heparin lyases, which are enzymes used for depolymerization of HS/heparin for sequence determination and production of therapeutic heparins. It was found that cleavage by lyase II is influenced by 3-OS, while digestion by lyase I is only affected by 2-OS. Lyase III exhibited sensitivity to both 3-OS and 2-OS.

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