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Irisa Trianti. Optimization of Recombinant Hemagglutinin Protein Production from Baculovirus Expression Vector System. Master's Degree(Biotechnology). King Mongkut's University of Technology Thonburi. KMUTT Library.. : King Mongkut's University of Technology Thonburi, 2552-12-27.
Optimization of Recombinant Hemagglutinin Protein Production from Baculovirus Expression Vector System
Name: Irisa Trianti
Abstract:
Hemagglutinin (HA), the major influenza viral surface glycoprotein, could
potentially be used as influenza vaccine. Baculovirus Expression Vector System
(BEVS) which has been known to produce high yield of recombinant protein with
biological activity, antigenicity and immunogenecity, the same as that of the natural
protein, was therefore employed to produce recombinant HA protein for this purpose.
The recombinant HA protein can be achieved after infection of genetically engineered
baculovirus harbouring HA gene into its insect cell host. High level of recombinant HA
protein production can be accomplished by optimization of several factors such as ratio
of cells and viruses (MOl), time of harvest (TOH), etc. It was found that high
recombinant HA protein production could be attained by infecting Sf-9 cells at the
density of lx106 cells/ml with recombinant HA baculovirus at MOl of 1. In addition,
recombinant HA protein production may be further optimized by reducing cell stress
caused by baculovirus infection using anti-stress agent. Therefore, supplementation of
anti-stress agents, catalase and selenium in the form of either sodium selenate
(Na2SeO4) or sodium selenite (Na2SeO3), into infected cell culture was proposed to
enhance the recombinant HA protein production. Result showed that recombinant HA
protein production was increased 1.3 times in comparison with that of the control at 3
days post infection when 0.625 ng/ml of catalase was supplemented into growth
medium. Moreover, when either Na2SeO3 or Na2SeO4 at the concentration of 0.4 and 0.2
ng/ml. respectively, was individually supplemented into growth medium, an increase in
recombinant HA protein produced of approximately 2.4 and 2.3 fold higher than that of
control at day 4 post infection, respectively, could be anticipated. It is worthy to note
that addition of these anti-stress agents not only increased level of recombinant HA
protein production but also shortened the production period from 5 days to 3 days (for
catalase) and 4 days (for selenium).
Effect of anti-stress agents on recombinant HA protein production was attained
through response surface methodology, Results showed that Na2SeO4 was considered
statistically significant (p<0.05), whereas effect of Na2SeO3 was subtle (p = 0.065).
Additionally, supplementation of catalase appeared to play insignificant role on
recombinant HA protein production. Further, it is interesting to note that interactions
among these anti-stress agents, i.e., Na2SeO3 x Na2SeO4, were all negative excluding
catalase x catalase, indicating their negative effects on recombinant HA protein
production. Result further demonstrated that optimal concentrations of Na2SeO3 and
Na2SeO4 leading to high recombinant HA protein production are, respectively, 0.18 and
0.37 ng/ml. Validation experiment conducted further showed that a slight increase in
recombinant HA protein production (1.3 folds higher than that of control) could be
anticipated on day 4 post infection providing the fact that, even on day 5 post infection,
level of recombinant HA protein produced was approximately 1.2 folds of that of
control which is better than that obtained when individual anti-stress agents was
supplemented into the growth medium.
King Mongkut's University of Technology Thonburi. KMUTT Library.
Address:
BANGKOK
Email:
info.lib@mail.kmutt.ac.th
Name: Kanokwan Poomputsa
Role:
Advisor
Created:
2552-12-27
Modified:
2552-12-27
Issued:
2009-12-27
วิทยานิพนธ์/Thesis
application/pdf
CallNumber:
BIT588
eng
DegreeName: Master of Engineering
Level: Master's Degree
Descipline: Biotechnology
©copyrights King Mongkut's University of Technology Thonburi
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0.027804 วินาที
Irisa Trianti
| Title | Contributor | Type |
|---|---|---|
| Optimization of Recombinant Hemagglutinin Protein Production from Baculovirus Expression Vector System
มหาวิทยาลัยเทคโนโลยีพระจอมเกล้าธนบุรี Irisa Trianti | Kanokwan Poomputsa | วิทยานิพนธ์/Thesis |
| Baculovirus surface display as antigen carrier for antibody production
มหาวิทยาลัยเทคโนโลยีพระจอมเกล้าธนบุรี Irisa Trianti | Kanokwan Poomputsa | วิทยานิพนธ์/Thesis |
Kanokwan Poomputsa