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Baculovirus Transfer Vectors

Although the flashBAC™ system is compatible with all transfer plasmids based on homologous recombination OET has developed a range of transfer plasmids that are designed to help you to get the most out flashBAC™. The pOET1 and pOET2 plasmids are designed to allow simple, easy cloning into the baculovirus system. The pOET3 and pOET4 plasmids are designed for improved expression of glycosylated, secreted and membrane-targeted proteins and are an ideal companion to the flashBAC™, flashBACGOLD™, and flashBACULTRA™ products. pOET5 is a dual promoter baculovirus transfer vector designed for high level expression of two foreign genes simultaneously. The vector is smaller than other available transfer vectors (4590bp) which greatly facilitate the cloning steps.

Transfer Vector Promoter Promoter Character Application
pOET1 or
pOET2
polh strong
expression
higher yield
pOET3 or
pOET4
P6.9 early
expression
better post-translational
modifications
pOET1N-His or
pOET1C-His
polh strong
expression
His tag at N or C terminus
pOET2N-His or
pOET2C-His
polh strong
expression
His tag at N or C terminus
pOET5 polh and P10 strong dual
expression
Express multi subunit proteins or
two proteins at the same time


pOET1

pOET1 is a baculovirus transfer vector designed for high level expression of foreign genes under the powerful AcMNPV polyhedron (polh) promoter. The vector is smaller than other available transfer vectors (4541bp) which greatly facilitate the cloning steps. It has a bacterial origin of replication and an ampicillin resistance gene for selection in E. coli. The polh sequences have been replaced by a multiple cloning site containing 14 unique restriction sites for insertion of the foreign gene in the correct orientation. The AcMNPV equences flanking the gene in the transfer vectors MCS allow recombination with the viral DNA to insert the expression cassette into the polh locus. pOET1 is compatible with any baculovirus system that utilizes homologous recombination in insect cells.

pOET1 Information and Vector Map (PDF)
pOET1N-His Information and Vector Map (PDF)
pOET1C-His Information and Vector Map (PDF)

pOET2

pOET2 is a baculovirus transfer vector designed for high level expression of foreign genes under the powerful AcMNPV polyhedron (polh) promoter. The vector is smaller than other available transfer vectors (4547bp) which greatly facilitate the cloning steps. It has a bacterial origin of replication and an ampicillin resistance gene for selection in E. coli. The polh sequences have been replaced by a multiple cloning site in the reverse orientation to pOET1, containing unique restriction sites for insertion of the foreign gene. The AcMNPV sequences flanking the gene in the transfer vectors MCS allow recombination with the viral DNA to insert the expression cassette into the polh locus. pOET2 is compatible with any baculovirus system that utilizes homologous recombination in insect cells.

pOET2 Information and Vector Map (PDF)
pOET2N-His Information and Vector Map (PDF)
pOET2C-His Information and Vector Map (PDF)

pOET3

pOET3 is a baculovirus transfer vector designed for high level expression of foreign genes under the late AcMNPV basic (P6.9) promoter. Using this promoter will provide earlier expression compared to the polyhedrin promoter. This has been shown to be beneficial when expressing proteins which require extensive posttranslational modifications i.e. glycosylation. The vector is smaller than other available transfer vectors (4530bp) which greatly facilitate the cloning steps. It has a bacterial origin of replication and an ampicillin resistance gene for selection in E. coli. The AcMNPV sequences flanking the gene in the transfer vectors MCS allow recombination with the viral DNA to insert the expression cassette into the polyhedrin locus. The polyhedrin sequences have been replaced by a multiple cloning site containing unique restriction sites for insertion of the foreign gene in the correct orientation. pOET3 is compatible with any baculovirus system that utilizes homologous recombination in insect cells.

pOET3 Information and Vector Map (PDF)

pOET4

pOET4 is a baculovirus transfer vector designed for high level expression of foreign genes under the late AcMNPV basic (P6.9) promoter. Using this promoter will provide earlier expression compared to the polyhedrin promoter. This has been shown to be beneficial when expressing proteins which require extensive post-translational modifications i.e. glycosylation. The vector is smaller than other available transfer vectors (4536bp) which greatly facilitate the cloning steps. It has a bacterial origin of replication and an ampicillin resistance gene for selection in E. coli. The AcMNPV sequences flanking the gene in the transfer vectors MCS allow recombination with the viral DNA to insert the expression cassette into the polyhedrin locus. The polyhedrin sequences have been replaced by a multiple cloning site containing unique restriction sites for insertion of the foreign gene in the correct orientation. pOET4 is compatible with any baculovirus system that utilizes homologous recombination in insect cells.

pOET4 Information and Vector Map (PDF)

pOET5

pOET5 is a dual promoter baculovirus transfer vector designed for high level expression of two foreign genes simultaneously under the powerful AcMNPV polyhedron (polh) promoter and the very late P10 promoter. The promoters are in opposite orientations to minimize recombination. The vector is smaller than other available transfer vectors (4590bp) which greatly facilitate the cloning steps. It has a bacterial origin of replication and an ampicillin resistance gene for selection in E. coli. The polh sequences have been replaced by two multiple cloning sites containing unique restriction sites for insertion of the foreign genes in the correct orientation. The AcMNPV sequences flanking the gene in the transfer vectors MCS allow recombination with the viral DNA to insert the expression cassette into the polh locus. pOET5 is compatible with any baculovirus system that utilizes homologous recombination in insect cells.

pOET5 Information and Vector Map (PDF)

pOET Sequencing Primers

pOET Sequencing primers are used to sequence inserts cloned into the multiple cloning site of any of the pOET transfer vectors. The forward primer anneals to the lef2 region upstream of the cloning site and the reverse primer anneals to the ORF1629 region downstream of cloning site.

The pOET sequencing primers anneal to the upstream and downstream regions of the multiple cloning site. They can be used to sequence across the MCS or amplify an insert within it by PCR. These primers are single-stranded oligonucleotides with 5'-hydroxyl and 3'-hydroxyl ends. All primers are supplied as 10 µM aqueous solutions.

pOET Sequencing Primers Information (PDF)


Ligation Independent Vectors (LIC Vectors)


LIC pOET1N_His transfer plasmid

LIC_pOET1N_His is a ligation independent cloning vector designed for the use in high throughput construct preparation and expression of genes under the powerful AcMNPV polyhedron (polh) promoter. Baculovirus transfer vector with His6 tag in 22-aa N-terminal fusion peptide, with TEV protease cleavage site. Includes sites for LIC cloning, and a “stuffer” fragment that includes the SacB gene, allowing negative selection of transformed bacteria on 5% sucrose.

LIC pOET1N_His transfer plasmid Information and Vector Map (PDF)

LIC pOET1N_His_GP64 transfer plasmid

LIC_pOET1N_His_GP64 is a ligation independent cloning vector designed for the use in high throughput construct preparation and expression of genes under the powerful AcMNPV polyhedron (polh) promoter. Baculovirus transfer vector with His6 tag and gp64 signal sequence in 44-aa N-terminal fusion peptide, with TEV protease cleavage site. Includes sites for LIC cloning, and a “stuffer” fragment that includes the SacB gene, allowing negative selection of transformed bacteria on 5% sucrose

LIC pOET1N_His_GP64 transfer plasmid Information and Vector Map (PDF)

LIC pOET3N_His transfer plasmid

LIC_pOET3N_His is a ligation independent cloning vector designed for the use in high throughput construct preparation and expression of genes under the late AcMNPV basic (p6.9) promoter. Baculovirus transfer vector with His6 tag in 22-aa N-terminal fusion peptide, with TEV protease cleavage site. Includes sites for LIC cloning, and a “stuffer” fragment that includes the SacB gene, allowing negative selection of transformed bacteria on 5% sucrose

LIC pOET3N_His transfer plasmid Information and Vector Map (PDF)

LIC pOET3N_His_GP64 transfer plasmid

LIC_pOET3N_His_GP64 is a ligation independent cloning vector designed for the use in high throughput construct preparation and expression of genes under the late AcMNPV basic (p6.9) promoter. Baculovirus transfer vector with His6 tag and gp64 signal sequence in 44-aa N-terminal fusion peptide, with TEV protease cleavage site. Includes sites for LIC cloning, and a “stuffer” fragment that includes the SacB gene, allowing negative selection of transformed bacteria on 5% sucrose

LIC pOET3N_His_GP64 transfer plasmid Information and Vector Map (PDF)