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Expression System
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Bacground
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Insect Cell Lines
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Vectors in Use
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Related Procedures
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1. Baculovirus Expression System
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Dr.
Danieli's lecture notes.
Invitrogen's
Bac to Bac manual (PDF).
Baculovirus
Expression System with Gateway Technology
The Baculovirus
tutorial (part of 72682 course material).
Guide
to Baculovirus Expression Vector systems (BEVS) and Insect Cell Culture
Techniques
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DH10Bac
Lepidopteran cells: general, SF9 (PDF),
SF21,
High-5,
Mimic-SF9 (PDF),
Baculovirus can infect Drosophila cells, however the virus will
not replicate in the infected cells and will not be able to infect subsequent
cells. No high-titer stocks can be generated from Drosophila cells.
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Gateway
destination vectors for baculovirus expression system (PDF), list
General vectors:
- FlashBac
- pAcGP67-C Baculovirus transfer vector
Bac-To-Bac vectors:
- MCS of PFastBac HC-1
- pFastBac1 - map, MCS
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Cryopreservation of SF9/H5 cells
Insect cells cryopreservation (S9, Sf21, S2, Tni)
Rapid thaw of SF9 and H5 cells
Transfection of Bacmid DNA into SF9 cells using ESCORT reagent
Transfection of FlashBac DNA & transfect vector into SF9 cells using ESCORT reagent.
Viral plaque assay
Viral plaque assay, abridged
Amplification of purified plaques.
Boosting protein expression in High5 & SF9 cells
Selenomethionine incorporation
Small scale purification of secreted His-tagged protein from conditioned medium
See also: Additional
resources
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| 2. InsectSelect Stable
Expression System |
InsectSelect
- pMIB/V5-His
InsectSelect
- pIB/V5 -His
InsectSelect
- pIZT/V5 -His
InsectSelect
- pIZ/V5 - HIS
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The InsectSelect System uses the OpIE2 promoter from the
Orgyia pseudotsugata multicapsid nucleopolyhedrosis virus for high-level
expression. This promoter is active in many common insect cell types including
SF9,
SF21
,
High
Five™, Mimic-SF9,S2,
MG1and KC1 cells. |
Invitrogen
vector search - type insectselect
pMIB/V5-His (map,
manual)
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See: Additional
resources |
| 3. DES: Drosophila
Stable
Expression System |
Drosophila
Expression System For the stable expression and purification of heterologous
proteins in Schneider S2 cells.
The Drosophila Expression System. |
S2
Schneider cells
D.Mel-2
cells (protocols)
These cells were isolated from late-stage Drosphila melanogaster
embryos and adapted to suspension growth in serum free medium.
The drosophila MT promoter used in this system does not work well in
lepidopteran cells (Sf9, sF21, and High Five).
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Invitrogen
vector search - type pmt or pac.
pMT/BiP/V5-His (map,
manual)
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See: Additional
resources |
