Cloning of the Glycoprotein 58 Region of the Guinea Pig Cytomegalovirus Glycoprotein B Gene
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Authors
Ebbing, Cathi
Issue Date
1997
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
Human cytomegalovirus (CMV) can cause deafness, blindness and varying
degrees of mental retardation in prenatally infected infants. CMV infections also
account for many medical complications such as pneumonia faced by individuals
whose immune systems have been weakened by chemotherapy or organ
transplants. Development of a vaccine to counter this virus will stem from the
discovery of structural proteins that will act effectively as antigens, thus eliciting
a humoral response from the human immune system. One CMV envelope protein,
glycoprotein B, has been determined to be the major target for virus-neutralizing
antibodies. Since guinea pig CMV undergoes transplacental and intrauterine
infection of the fetus in a manner similar to humans, guinea pig CMV was used in
this experiment as an animal model. Cloning and expression of the carboxy I
terminus of guinea pig gB, (identified as gp58) was undertaken as a preliminary
step towards future examination of the antigenic potential of gp58. Attempts
were made, using polymerase chain reaction and other standard subcloning
techniques, to clone gp58 and to ligate it into New England Biolab's pMal-p2
expression vector for eventual expression and purification using the Protein
Fusion and Purification System. A PCR generated fragment was successfully
ligated into pMal-p2, however it was in the incorrect orientation for proper
protein translation. Restriction endonuclease digestion of a plasmid previously
engineered to contain the gp58 segment was performed. The resulting fragment
was then ligated into the vector. Subsequent analysis of this fragment revealed
that the ligated insert was not· the desired gp58 region of the guinea pig
cytomegalovirus.
Description
vi, 21 p.
Citation
Publisher
Kalamazoo College
License
U.S. copyright laws protect this material. Commercial use or distribution of this material is not permitted without prior written permission of the copyright holder.