Download 61 - Lab Times

6-2011
Products
Lab Times
page 61
Product survey: Expression vectors
Gene Delivery Boys
In the early seventies Herb Boyer, Stanley Cohen and Annie Chang published two groundbreaking papers on restriction
enzyme-based DNA recombination and expression of Staphylococcus aureus genes in E. coli from “hybrid plasmids”.
Today almost every group keeps expression vectors in the freezer to easily express their favourite genes.
H
erb Boyer and Stanley Cohen opted for E. coli as a host for their early expression plasmids. A decision
that approved pretty clever and has passed
the test of time. Life scientists developed
many other expression systems since Boyer’s and Cohen’s pioneering work, ranging from different Bacillus species, yeasts, insect cells, silkworm larvae, chloroplasts to
mammalian cells. However,
E. coli is still the number
one expression system in
most labs.
According to a recent
survey, 60% of recombinant
gene products are currently expressed in E. coli cells (Hans Peter Sørensen, Microbial Cell Factories 2010, 9:27). That’s for good reasons:
expression of heterologous genes in E. coli is
cheap, easy and fast. Besides that, countless
expression vectors suitable for E. coli cells
are available, equipped with all kinds of
promoters, selection markers, tags and fusion proteins that molecular biologists can
think of. Typical promoters are, e.g., IPTG
inducible lac promoters, trp and phoA promoters induced by tryptophan and phosphate depletion, respectively, and araB promoters that are induced by arabinose.
Popular system
T7 promoters derived from the bacteriophage T7 are also very common and a
crucial part of the popular pET expression
system developed by T7-RNA expert William Studier in the late eighties. The basic idea of Studier’s pET system is pretty
straightforward. Since T7 RNA polymerase
is highly specific for the T7 promoter, background expression of genes cloned in a pET
expression vector downstream of a T7 promoter is negligible in E. coli having no T7
RNA polymerase. Hence, to kick-start the
transcription, T7 RNA polymerase must be
co-expressed in E. coli cells in parallel. This
is usually done by integrating the T7 RNA
polymerase gene into the E. coli chromosome together with an upstream T7 pro-
moter or by transfecting the cells with the
phage lCE6, which produces T7 RNA polymerase.
E. coli does not, however, express every recombinant gene as a soluble and functional protein. Some end up as clotted, unfeasible inclusion bodies inside the cell.
Another shortcoming is its inability to perform typical eukaryotic post-translational modifications, such as glycosylation, phosphorylation
or acetylation. If
you spend most
of your time trying to get inclusion bodies into
solution or worrying about expressed proteins lacking
post-translational modification, then
it’s high time to say good-bye to E. coli and
check out other expression hosts.
Pichia takes the lead
Life scientists usually try yeast cells
capable of doing many post-translational modifications as an easy and fast alternative, before establishing a laborious and
time consuming mammalian expression
system. You may probably not guess, however, which yeast expression system they
choose most frequently. No, it’s not the baker’s yeast Saccharomyces cerevisiae, it’s the
methylotropic yeast Pichia pastoris! Almost
20% of today’s recombinant proteins are
expressed in Pichia pastoris cells.
Pichia pastoris is an unusual type of
yeast that does not grow on glucose as a
preferred carbon source like Saccharomyces but on methanol. The first step in methanol utilisation requires oxidation of methanol to formaldehyde by the enzyme alcohol oxidase (AOX). Since Pichia needs large
amounts of the AOX protein to cope with
this task, it developed an extremely strong,
methanol-inducible AOX1 promoter that
propels the transcription of the AOX gene.
Molecular biologists took advantage of the
powerful AOX1 promoter and cloned it upstream of recombinant genes into suitable
Pichia pastoris expression vectors. Induction of the promoter by methanol leads to
an almost exclusive production of the recombinant protein. Usually, these vectors
contain further elements, which facilitate
cloning or encode, e.g., secretion signals
that guide the expressed protein out of the
cells into the media.
Smart expression platform
In a recent PLoS ONE paper, Hakan Dortay and his colleagues from the Institute of
Biochemistry and Biology at the University of Potsdam describe an interesting protein expression platform, based on E.coli,
Pichia pastoris and the alternative hosts
Klyveromycis lactis and Leishmania tarentolae (completed by an in vitro transcription/
translation system), that allows fast and
easy expression of a wide spectrum of proteins (Dortay et al., PLoS ONE 6(4): e18900.
doi:10.1371/journal.pone.0018900). To express the proteins in the aforementioned
hosts, the group constructed ten expression
vectors by a ligation-independent cloning
(LIC) technique.
LIC is based on the 3’-5’-exonuclease
activity of T4 DNA polymerase that creates single stranded 5’-tails in DNA fragments, e.g., PCR products and complementary overhangs in LIC vectors. After mixing
LIC vectors and PCR amplicons, the complementary cohesive ends are annealed. To
construct N- or C-terminal fusions, Dortay
and his colleagues designed two PCR fragments per vector, one with and one without stop codon, and inserted them into LICcompatible expression vectors.
Since the LIC efficiency is more or less
100% (according to the authors) and the
insertion of the fragments into the LIC vectors can be done in parallel, the LIC-based
expression platform permits rapid parallel
protein expression in four of the most popular and reliable hosts. That should be sufficient for a great deal of protein expression
projects. If not for yours, then take a closer
look at the next pages for additional expression vectors and systems.
Harald Zähringer
page 62
Lab Times
6-2011
Products
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
Active Motif
La Hulpe, Belgium
www.activemotif.com
Contact: Jörg Plümpe
Phone +32 2653-0001
[email protected]
Agilent Technologies
www.genomics.agilent.com
Contact: Mimi Estey
Phone +1 858-373-6358
[email protected]
Amsbio
Abingdon, UK
www.amsbio.com
Contact:
Phone +44 1235-828200
[email protected]
Miscellaneous, Specialities,
Generally
Price
CMV promoter and kanamycin resistance marker —
Obtain different functionalities by exchanging the
LigandLink label — No changes of the characteristics
of your protein of interest — Pre-validated expression
systems for p53, STAT1 und NF-kB p65 available
On request
[EUR]
LigandLink Expression system
(LigandLink
pLL-1kit)
Eukaryotic cells
System to express and label proteins
within living cells — Protein
localisation and trafficking —
Monitor protein: protein interactions
— Protein capture
StrataClone
Mammalian
Untagged
Mammalian cells
High-level protein expression —
Untagged, CMV, BGH polyA, neo/kan
Rapid cloning of blunt-ended PCR
products in topoisomerase-activated
vector arms
394.(20 rxn)
StrataClone
Mammalian
N-Flag
Mammalian cells
High-level protein expression of
N-terminal FLAG-tag, CMV, BGH-polyA, neo/kan
epitop-tagged proteins — Rapid
cloning of blunt-ended PCR products
394.(20 rxn)
StrataClone
Mammalian
C-Flag
Mammalian cells
High-level protein expression of
C-terminal FLAG tag, CMV, BGH-polyA, neo/kan
epitop-tagged proteins — Rapid
cloning of blunt-ended PCR products
394.(20 rxn)
StrataClone
Mammalian
N-cMyc
Mammalian cells
High-level protein expression of
N-terminal cMyc-tag, CMV, BGH-polyA, neo/kan
epitop-tagged proteins — Rapid
cloning of blunt-ended PCR products
394.(20 rxn)
StrataClone
Mammalian
C-cMyc
Mammalian cells
High-level protein expression of
C-terminal cMyc-tag, CMV, BGH-polyA, neo/kan
epitop-tagged proteins — Rapid
cloning of blunt-ended PCR products
394.(20 rxn)
LacSwitch II In- Mammalian cells
ducible Mammalian Expression
System
Inducible mammalian expression
system that utilises an improved
vector system
Tight repression — Rapid induction (4-8 h)
823.-
Complete
Mammalian cells
Control Inducible
Mammalian Expression System
Inducible mammalian expression
system (plasmid & retroviral based)
Tightest repression — High induction rate — Plasmid
based, retroviral based, Ecdysone-inducible vector
896.418.416.-
pCMV-Tag
vectors
Mammalian cells
Series of epitope tagging
mammalian expression vectors
cMyc/FLAG-tag, CMV, SV40-polyA, neo/kan — Price
depending on vector type
414.- to
560.-
pCMV-3Tag
vectors
Mammalian cells
Series of epitope tagging
mammalian expression vectors
featuring three copies of the epitope
tag in a variety of configurations
cMyc/FLAG-tag, CMV, SV40-polyA, neo/kan or hyg —
Price depending on vector type
424.- to
560.-
pSG5 Expression Mammalian cells
Vector
For in vitro and in vivo expression
High copy number — Transient expression
424.-
pBK-CMV Vector
Mammalian and
bacterial cells
For expression in mammalian and
bacterial systems
CMV, SV40-polyA, neo/kan, lac promoter
456.-
PrecisionShuttle
Mammalian
Vectors
Mammalian
Available with different tags (myc,
His, GFP...) — Easy subcloning of
ORF from one vector to another —
CMV promoter
Tags: Myc, DDK, His, HA, FC — Fluorophores: GFP, TurboGFP, TurboRFP, TurboYFP, TurboFP602, TurboFP635,
mKATE, mGFP, mRFP, mYFP, mBFP, mCFP — Selection:
neomycin, blasticidin, hygromycin, puromycin
On request
pCMV6-Neo,
Kan/Neo
Mammalian
CMV promoter, for establishment of
stable cell lines
Selection: neomycin
On request
pCMV6-XL 4,
5, 6
Mammalian
CMV promoter, transient
transfection only
--
On request
Vector for miRNA Mammalian
expression,
pCMV-MIR
Vector for miRNA expression driven
from CMV promoter with GFP as
control — Can be used for
establishment of stable cell lines
Overexpression of miRNA — GFP to monitor
transfection efficiency — Neomycin selection marker
On request
pTune
Inducible vector that tightly regulates >99% repression — IPTG induction — Tags: DDK and
gene expression — Expression is
Myc — Neomycin selection marker
controlled by repressor proteins and
RNAi target sequence — Compatible with PrecisionShuttle system
On request
Vector for bacterial expression
with His and/or GST tag —
IPTG induction
On request
Mammalian
PrecisionShuttle Bacterial
Bacterial Vectors
Tags: His and GST — Selection: Ampicillin
6-2011
Products
Lab Times
page 63
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
Amsbio (continued)
Heidelberg, Germany
www.biocat.com
Contact: Elke Gamer
Phone +49 6221-7141516
[email protected]
Biomol
Hamburg, Germany
www.biomol.de
Contact: Edgar Lipsius
Phone +49 40-853260-37
[EUR]
Mammalian
CMV-based vectors for high expres- Tag: HA — Selection: Neomycin
sion — Transient & stable transfec
tion — convenient multiple cloning site
On request
gWiz Vectors
Mammalian
High expression vectors for both
in vitro and in vivo expression with
reporter genes
Reporter genes: GFP, luciferase, ß-galactosidase,
chloramphenicol acetyltransferase (CAT), secreted
alkaline phosphatase (SEAP)
On request
Lentivector
Mammalian
Lentiviral expression vectors for
cDNA and shRNA
GFP, RFP for transduction control —
Selection: puromycin, blasticidin, neomycin
On request
PepIdent
E.coli
Hosting and expression of customdesigned deterministic bio-peptide
libraries derived from entire proteins
or specific domains etc. —
Libraries are designed with a
proprietary design tool and hosted
as DNA on plasmids
Antibody screening — Epitope screening
On request
ACEMBL series
E.coli, insect cells 3 different systems (5 vectors each) Recombineering — Protein production and tracking
(Sf9, Sf21, High- for multi-protein and protein complex
expression/ production
Five, potentially
S2), mammalian
cells
Toggle ACEMBL
Any organism
that is amenable
to plasmid-based
modification;
focus on microorganismal production strains
Merzhausen, Germany
www.atg-biosynthetics.com
Contact: Hubert Bernauer,
Alexander Craig
Phone +49 761-8889424
[email protected]
BioCat
Price
phCMV
Contact: see page 62
ATG:biosynthetics
Miscellaneous, Specialities,
Generally
On request
Flexible generation of complex
multi-gene constructs, e.g. for
(combinatorial) protein complexes
and pathway designs
Complex DNA assemblies — Pathway design
On request
Expresso Cloning E.coli
and Expression
System
Rapid directional cloning using
Expressioneering technology
Expression of N- or C-terminal 6xHis-tagged proteins —
Vector available with tunable rhamnose or strong
T7 promoter — Cleavable SUMO tag version available
to improve protein solubility — Effortless, enzyme-free
cloning into ready-to-use Expresso vectors —
High efficiency: >90% recombinants
235.-/399.(5/10 rxn)
299.- /499.(5/10 rxn,
SUMO Solubility Tag)
cDNA Cloning
and Expression
Lentivectors
Efficient expression of the gene of
interest in most mammalian cell
types
Strong and ubiquitous expression of the gene of interest Depending
on vector
— Different promoters available: CMV, MSCV, EF1alpha, PGK, UbC — Choice of fluorescent or selection
marker — Efficient delivery to most cell types —
Self-inactivating, replication-incompetent HIV-based
third generation lentivector
Cumate Switch
Mammalian cells
Inducible Exincluding hardpression System to-transfect cells
like primary cells
and non-dividing
cells
Inducible expression of the gene of
interest in mammalian cells
Cumate-based inducible expression — HIV-based
SparQ Cumate Switch lentivectors — Zero background
leakiness — Robust induction and dynamic titratability — Turn on -> off -> on again — Induction can be
tracked with co-expressed GFP or RFP
Depending
on vector
iLenti siRNA Expression Vectors
Mammalian cells
including hardto-transfect cells
like primary cells
and non-dividing
cells
Lentivector-based gene silencing in
mammalian cells
Efficient gene knockdown using 27-29 bp siRNA
expressed from convergent promoters — Hairpin
loop structures avoided for easier propagation and
sequencing — EGFP marks siRNA expressing cells
— Replication-incompetent HIV-based third generation
lentivector — Same price for premade and custom
cloned constructs
150.(500 ng)
440.(4 x 500 ng)
PiggyBac Transposon System
Human, mouse
and rat cells
Generation of transgenic cell lines
by transposition with only one
transfection
Effective in human, mouse and rat cells — No cargo
limit: integrate 10-100 kb — Reversible integration —
Also available as cumate-based inducible vector
Depending
on vector
Minicircle Vector
Technology
Mammalian cells
(dividing and
non-dividing)
Prolonged expression of a transgene
without integration into the genome
Episomal DNA devoid of any bacterial plasmid backbone Depending
— Sustained expression over weeks — No integration into on vector
the genome — Works in vitro & in vivo — Vector formats
with different promoters & fluorescent reporters available
Complete
RetroMax
System
Mammalian cells
Retroviral expression system for
Contains 4 expression vectors with different cloning
transient or stable protein expression sites and promoters — Provides 3 packaging vectors
for the infection of diverse host target cells —
High retrovirus titer (2-5 x 106 CFU/ml in 293 cells) —
Retroviral constructs are safety-modified
Mammalian cells
including hardto-transfect cells
like primary cells
and non-dividing
cells
535.- (1 kit)
page 64
Lab Times
6-2011
Products
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
IBA
Göttingen, Germany
www.iba-go.com
Contact: Isabel Schuchardt
Phone +49 551-50672-0
[email protected]
Jena Bioscience
Jena, Germany
www.jenabioscience.com
Contact: Reinhard Breitling
Phone +49 3641-6285125
reinhard.breitling@
jenabioscience.com
Merck Millipore
Miscellaneous, Specialities,
Generally
Price
[EUR]
Classic Cloning
Vectors
pASK-IBA /
pPR-IBA
E.coli
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag)
proteins under control of the tet
promoter (pASK) or T7 promoter
(pPR) in E.coli
For single N-/C-terminal fusions as well as double-tags
in all possible combinations — Protease cleavage of
the Tag possible — Tightly regulated expression with
tet promoter — pASK-IBA vectors with Ampicillin or
Chloramphenicol resistance available —
For Cytosolic or periplasmic expression
155.- (5 μg)
Classic Cloning
Vectors
pEXPR-IBA
Mammalian cells
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag)
proteins under control of the CMV
promoter in mammalian cells
For single N-/C-terminal fusions as well as double-tags
in all possible combinations — Protease cleavage of
the tag possible — Secretion of proteins possible
From 185.(5 μg)
StarGate Vectors E.coli
pASG-IBA /
pPSG-IBA
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag,
GST-tag and FLAG-tagged) proteins
under control of the tet promoter
(pASG) or T7 promoter (pPSG) in
E.coli
Simple and fast cloning procedure (StarGate Cloning)
— Easy screening for best expression results —
Multitude of N-/C-terminal fusion tags available as
well as double tags in all possible combinations —
Tightly regulated expression with tet promoter —
For cytosolic or periplasmic expression
80.- (5 rxn)
StarGate Vectors Mammalian cells
pESG-IBA/pCSG
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag,
GST-tag and FLAG-tagged) proteins
under control of the CMV promoter in
mammalian cells
Simple and fast cloning procedure (StarGate Cloning)
— Easy screening for the best expression results —
Multitude of N-/C-terminal fusion tags available as
well as double tags in all possible combinations —
Secretion of proteins possible — Transient (pESG)
and episomal (pCSG) vectors available
80.- (5 rxn)
StarGate Vectors Yeast SaccharopYSG-IBA
myces cerevisiae
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag,
GST-tag and FLAG-tagged) proteins
under control of the CUP1 promoter
in yeast cells
Simple and fast cloning procedure (StarGate Cloning)
— Easy screening for the best expression results —
Multitude of N-/C-terminal fusion tags available as
well as double tags in all possible combinations —
Regulated expression due to inducible CUP1 promoter
80.- (5 rxn)
StarGate Vectors Insect cell lines
pLSG-IBA
Expression of affinity tagged (Streptag, One-Strep-tag, 6xHistidine-tag,
GST-tag and FLAG-tagged) proteins
under control of the polyhedrin
promoter in insect cell lines
Simple and fast cloning procedure (StarGate Cloning)
— Easy screening for the best expression results —
Multitude of N-/C-terminal fusion tags available as well
as double tags in all possible combinations
80.- (5 rxn)
LEXSY protein
expression kit
EGE-1300
Leishmania
tarentolae
Constitutive protein expression in
unicellular eukaryotic host
Fast growth — High yields —
Mammalian PTMs — Easy purification
960.-
LEXSY protein
expression kit
EGE-1410
Leishmania
tarentolae
Inducible protein expression in
unicellular eukaryotic host
Fast growth — High yields —
Mammalian PTMs — Easy purification
2,250.-
Cloning and expression of recombinant proteins in E.coli under tight
control of strong bacteriophage T7
transcription and (optionally)
translation signals
Selection of vectors and host strains to control basal
and induced expression levels — Large choice of
N-teminal and C-terminal fusion tags — Specialised
vectors and hosts — Rapid recombination cloning to
prepare pEXPR clones (Gateway pDEST vectors)
See website
Complete system for rapid plasmidbased expression in insect cells
Provides most rapid expression compared to baculovirus-based system — Insect cells are transfected with
expression plasmid containing hr5 enhancer and early
promoters — Useful for rapid, high-level expression
and screening — Can also be used to produce stably
transfected cell lines
See website
Dual-purpose vectors for direct
transfection or baculovirus
production
Compatible with both InsectDirect transfection for plas- See website
mid-mediated expression or to generate recombinant baculovirus — Single construct can be used for both rapid HT
screening and optimised, high-yield protein expression —
AcNPV hr5/ie1 promoter/enhancer for plasmid-mediated
and early baculovirus expression — AcNPV p10 very late
promoter for late baculovirus expression
Nonlytic expression in insect cells
plus tightly regulated E.coli
expression
Designed for rapid expression of target genes in both
E.coli and insect cells — Features tightly controlled
T7lac promoter for protein expression in E.coli —
Features hr5 enhancer and ie1 promoter for plasmiddirected target expression screening in insect cells
See website
Multisystem expression vectors — Optimised transcription and translation signals for E.coli, baculovirus, and
mammalian expression (T7lac promoter, p10 promoter
and either beta-actin or CMV promoter, respectively)
See website
pET vector series E.coli (DE3)
strains providing
a source of T7
RNA polymerase
A division of Merck
www.millipore.com
Contact: technischerservice@
merckgroup.com
Phone:
+49 6196-494-299 (DE)
InsectDirect
+41 433994049 (CH)
system
+43 820874464 (AT)
Sf9 insect cells
pIEx/Bac vectors Sf9 insect cells
pBiEx vectors
E.coli (DE3)
strains or Sf9
insect cells
pTriEx vectors
One construct for efficient expression
E.coli (DE3)
strains, Sf9 insect in bacterial, insect and mammalian
systems
cells and mammalian systems
6-2011
Products
Lab Times
page 65
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
MoBiTec
Göttingen, Germany
www.mobitec.com
Contact: Arne Schulz
Phone +49 551-70722-0
[email protected]
PAA Laboratories
Pasching, Austria
www.paa.com
Contact: Anne Schönbauer
Phone +49 6421-17539-0
[email protected]
PlasmidFactory
Bielefeld, Germany
www.PlasmidFactory.com
Contact: Martin Schleef
Phone +49 521-2997350
[email protected]
5 Prime
[EUR]
Bacillus megaterium, Strain
WH320 —
Bacillus megaterium, Strain
MS941
High yield protein expression —
Vectors for intracellular expression
or secretion
High performance shuttle-vectors w. optimised sequence
— Protein yield up to 10 times enhanced compared to
protein expression with basic plasmid — Encoding C- or
N-terminal His-tag for versatile purification — Secretion
with LipA or YocH signal peptide up to 9-fold increased
— Protoplasts ready for transformation available
970.- to
1,010.(Depending
on expression system)
NICE Expression
System for
Lactococcus
lactis
Lactococcus lactis, Strain NZ9000
— Lactococcus
lactis, Strain NZ3900, food grade
— E.coli, host
Strain MC1061
Nisin controlled gene expression system for the production of:
homologous and heterologous
proteins, prokaryotic and eukaryotic
membrane proteins, exo-polysaccharides, ingredients through genetic
engineering
Tightly controlled gene expression allows production of
toxic proteins — Secretion of proteins into the medium
— Less endogenous and no exogenous proteases,
no inclusion bodies — Endotoxin-free food grade
expression system — Simple fermentation, scale-up
and downstream processing
358.-
Bacillus subtilis
Expression
System
IPTG inducible protein production
For intracellular
expression: Bacil- — Intracellular and secretion —
lus subtilis, Strain Cold-inducible protein production
1012 wild type,
Bacillus subtilis, Strain 168
Marburg — For
secretion vectors:
Bacillus subtilis,
Strain WB800N
Bacillus subtilis is non-pathogenic and is considered
as a GRAS organism (generally regarded as safe) —
It is capable of secreting functional extracellular
proteins directly into the culture medium
353.-
CMV-based
constitutive Expression Vectors
E.coli
High level constitutive protein
expression
380.Maximised high-level expression with optimised CMV
promoter — G418 resistance gene for the selection of
stable cell lines — Optional N- or C-terminal HA fusion
tags — Extensive multiple cloning region for convenient
and easy cloning
pAA1, pAA 2
(Transfer Vectors)
Insect cells
Baculovirus expression
Small vector sizes for efficient transfection — Small
size for easy cloning — High level expression from the
polh promoter — Compatible with all baculovirus
systems based on homologous recombination
pAA3, pAA4
(Transfer Vectors)
Insect cells
Baculovirus expression
On request
High level expression for foreign genes under the late
AcMNPV basic (p6.9) promoter — Enhanced posttranslational protein processing — Optimised production of
complex proteins such as glycoproteins — Compatible
with all baculovirus systems based on homologous
recombination
Plasmid DNA
Minicircle DNA
Produced in
E.coli;
for expression in
mammalian cells
and tissue
Transfection, Gene therapy, Gene
vaccine, Reporter gene assays, Virus
production, e.g. AAV
Contract manufacturing — In Stock plasmids (AAV
helper and packaging plasmids, reporter genes,
S/MAR) — Patented minicircle technology —
Higher dose per mg — Production of minicircle DNA
free of bacterial sequences
PerfectPro
E.coli
High-level expression of N-terminally Multiple cloning sites and translation stop codon in all
429.6xHis-tagged proteins from the T5
three reading frames — Powerful phage T5 promoter
promoter
— Expression of short peptides as DHFR-fusion proteins
— Easy purification of N-terminally 6xHis-tagged proteins
E.coli
High-level expression of C-terminally
6xHis-tagged proteins from the T5
promoter
Full-length protein purification ensured by C-terminal
6xHis-tag — Powerful phage T5 promoter regulated by
double lac operator — Expression of short or poorly
expressed proteins as DHFR-fusion proteins —
Easy purification of C-terminally 6xHis-tagged proteins
PerfectPro 100
Double Tag
Vector DNA
E.coli
High-level expression of proteins
with an N-terminal 6xHis-tag and a
C-terminal Tag 100
207.C-terminal Tag 100 — Powerful phage T5 promoter
regulated by double lac operator — Easy purification of
N-terminally 6xHis-tagged proteins
PerfectPro 30Xa
Vector DNA
E.coli
High-level expression of N-terminally Removal of the 6xHis-tag peptide by Factor Xa Protease
6xHis-tagged proteins including a
cleavage site — Powerful phage T5 promoter regulated
Factor Xa Protease recognition site
by double lac operator — Easy purification of
N-terminally 6xHis-tagged proteins
pSEX81
E.coli XM-1 Blue
Phage Display — Antibody libraries
Phagemid cassette vector — Single-chain Fv –pIII fusion 260.-
pOPE101
E.coli XM-1 Blue
Phage Display — Antibody libraries
Phagemid cassette vector — Single-chain Fv –pIII fusion 260.-
Hyperphage
M13
Helperphage for Phage Display
Improves panning efficiency for high and low affinity
binders
C-6-His Vector
Set
Heidelberg, Germany
www.progen.de
Contact: Gerda Bruder
Phone +49 6221-8278-0
[email protected]
Price
hp-Vector
Expression
Systems for
Bacillus
megaterium
Hamburg, Germany
N-6-His Vector
www.5PRIME.com
Set
Contact: Ruth Rottscheidt
Phone +49 40-3197927-0
[email protected] PerfectPro
Progen Biotechnik
Miscellaneous, Specialities,
Generally
438.-
On request
On request
(depending
on product
and amount
produced)
408.-
204.-
475.-
page 66
Lab Times
6-2011
Products
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
Progen Biotechnik
(continued)
Contact: see page 65
Promega
Mannheim, Germany
www.promega.com
Contact: Michaela Mack
Phone +49 621-8501164
michaela.mack@
promega.com
PromoCell
Heidelberg, Germany
www.promokine.info
Contact: Technical Support
Phone +49 6221-64934-0
[email protected]
Miscellaneous, Specialities,
Generally
Price
[EUR]
pAc-kappa-CH3
Baculovirus
Single-chain antibody cloning
Phagemid cassette vector — Functional single-chain
antibody fragments
260.-
pAc-kappa-Fc
Baculovirus
Single-chain antibody cloning
Phagemid cassette vector — Functional single-chain
antibody fragments
260.-
pAc-lambda-CH3 Baculovirus
Single-chain antibody cloning
Phagemid cassette vector — Functional single-chain
antibody fragments
260.-
pAc-lambda-Fc
Baculovirus
Single-chain antibody cloning
Phagemid cassette vector — Functional single-chain
antibody fragments
260.-
HaloTag Fusion
(N-Terminal
or C-Terminal)
Mammalian
Expression Flexi
Vectors
Mammalian cells
E.coli
Variable expression of proteins in
Multifunctional tag for various applications —
mammalian cells — Cell imaging
Flexi vector for simple entry and transfer to other
of protein localisation or trafficking
Flexi vectors
in conjunction with the fluorescent
HaloTag Ligands — Purification from
the cells or protein pull-downs with
other proteins
Untagged Flexi
Mammalian Expression Vectors
Mammalian cells
High-level expression of proteins in
mammalian cells
Flexi vector for simple entry and transfer to other
Flexi vectors
pTargeT
Mammalian
Expression
Vector System
Mammalian cells
Constitutive expression of cloned
DNA inserts in mammalian cells
T cloning vector for direct insertion of PCR products
pCMVTnT Vector
Mammalian cells
Convenient expression of cloned
genes using both in vitro and in vivo
expression systems
SP6 and T7 polymerase promoters — Presence of
RNA phage promoters — 5´ ß-globin leader
sequence — CMV enhancer/promoter region —
ß-globin/IgG chimeric intron
pCI and pSI
Mammalian Expression Vector
Mammalian cells
Constitutive expression of cloned
DNA inserts in mammalian cells
A ß-globin/IgG chimeric intron located downstream
of the enhancer/promoter region can further increase
expression
Inducible
E.coli
T7-driven Flexi
Vectors for E.coli
Expression
Expression of proteins in E.coli from
the T7 RNA polymerase promoter
Flexi vector for simple entry and transfer to other Flexi
vectors — Available with GST, HQ and HaloTag
pPK-CMV Expression Vectors
Constitutive high-level gene
expression in mammalian cells
Suitable for both stable and transient expression —
Three variants available for expression of native and
N-terminal or C-terminal HA-tagged proteins — Small
plasmid size — Convenient multiple cloning site —
Positive control vector containing a CAT gene
269.(25 μg)
Extremely high constitutive expression of recombinant proteins in a
wide variety of mammalian cell types
— GFP or Luciferase can be fused
to the N- or C-terminal of the target
proteins or simply co-expressed with
these two reporter proteins — They
also enable creating GFP or Luciferase expressing stable cell lines
Convenient choice of GFP or Luciferase reporter genes
— Two multiple cloning sites — Kanamycin/Neomycin
resistance genes — SV40 polyadenylation sign
provides for stable mRNA — pUC origin of replication
yields high copy number of vector in E.coli
329.(20 μg)
StarGate Cloning Mammalian cells, Rapid, convenient and highly
Systems
E.coli, yeast cells, efficient sub-cloning of a target gene
that can be simultaneously fused
insect cells
with many different genetic
surroundings
One-tube, easy-to-handle sub-cloning —
Minimal modification of the gene of interest —
Improved, systematic screening of different elements
— Inherent highest level cloning efficiency —
Enables easy site-directed mutagenesis of target genes
295.- (Entry
Cloning Set)
249.- (Transfer Reag. Set)
79.- (Acceptor Vector)
429.- (Fusion
Cloning Set)
395.- (Newcomer Set)
GeneOFF shRNA
Vector Kits
Expression of shRNAs in many different cell types —
With U6- or H1 RNA Polymerase III-promotor (H1 variant
with or without GFP reporter gene) — Small plasmid
size for increased transfection efficiency — Suitable for
both stable and transient expression of shRNA — Fast
and easy cloning of the shRNA coding DNA sequence
219.(25 rxn)
249.(GFP reporter
variant,
25 rxn)
Mammalian cells
pPK-CMV Fusion Mammalian cells
Vectors
Mammalian cells
High-level expression of specific
shRNA molecules in mammalian
cells — shRNAs are subsequently
processed into functional siRNAs for
efficient suppression of target genes
(gene silencing, gene knockout, RNA
interference)
Please
contact your
local branch
or distributor
6-2011
Products
Lab Times
page 67
Expression Vectors
Company/Distributor Name of
Host cells
Applications
product
Sirion Biotech
Premade Adeno- HEK 293
virus for gene
Martinsried, Germany
overexpression
www.sirion-biotech.de
Contact: Kathrin Schmitt
Phone +49 89-700 9619913
Premade Adeno- HEK 293
[email protected]
virus for gene
silencing
Takara/Clontech
Takara Bio Europe
St.Germain-en-Laye, France
www.clontech.com
Contact:
Phone +33 1 3904 6880
(Europe)
Toll Free Phones
0800 1825 178 (DE)
0808 234 8063 (UK)
0800 563 629 (CH)
0800 296 141 (AT)
[email protected]
Miscellaneous, Specialities,
Generally
Price
[EUR]
670.Transgene overexpression into nearly Highly efficient gene delivery potential, even into
all eukaryotic cells
primary cells — Degree of overexpression adjustable by
using different virus amounts — Reproducible results
by high efficient titration
Efficient gene silencing into nearly all Highly efficient gene knockdown potential, even into
eukaryotic cells
primary cells — Pre-validated shRNA sequences
guarantee for knockdown ≥ 80% on mRNA —
Reproducible results by high efficient titration
750.-
Bicistronic IRES
Vectors
Mammalian cells
Simultaneous expression of two
separate proteins from one mRNA
transcript
Vectors with EF1alpha promoter (for robust expression
in hematopoietic or stem cells) or with CMV promoter
— Different antibiotic selection markers: puromycin,
hygromycin, G418 or bleomycin — Different fluorescent proteins
On request
Bidirectional
Promoter Vectors (pBI-CMV1
vectors)
Mammalian cells
Constitutive expression of two
proteins at similar levels
pBI-CMV1 vector allows the constitutive expression of
two proteins of interest — pBI-CMV vectors for
expression of one protein of interest and one reporter
protein (fluorescent protein or luciferase)
On request
Myc-Tag and
HA-Tag Vectors
Mammalian cells
Expression of a protein fused to
either the c-Myc or hemagglutinin
(HA) epitope tag
Co-immunoprecipitation of myc-tagged and HA-tagged
proteins in mammalian cells — Immunostaining,
western blot or ELISA with c-Myc and HA antibodies —
Vectors with EF1alpha promoter (for robust expression
in hematopoietic or stem cells) or with CMV promoter
On request
pBApo-vectors
Mammalian cells
Basic vectors for gene expression in
mammalian cells
Vectors with EF1alpha promoter (for robust expression
in hematopoietic or stem cells) or with CMV promoter
— Different antibiotic selection markers: puromycin
or G418
On request
ProteoTuner
Mammalian cells
Expression of a protein fused to
destabilization domain (DD)
Expressed protein is destabilised by DD domain —
Allows regulation of expressed protein on
translational level — Rapid kinetics: level of expressed
protein can be changed within minutes
On request
Fluorescent
protein vectors
Mammalian cells, Expression of a protein fused to
fluorescent proteins
E.coli
20 different fluorescent proteins: blue, green, yellow,
orange, red and far red — Vectors with EF1alpha
promoter (for robust expression in hematopoietic or
stem cells) or with CMV promoter — Vectors for
bacterial expression
On request
Tet-regulated
vectors
Mammalian cells
Inducible gene expression
Tet-On 3G for lowest background and highest sensitivity
— Vectors with EF1alpha promoter (for robust
expression in hematopoietic or stem cells) or with
CMV promoter — Vectors with fluorescent proteins —
Tet Express for fast set up and fast induction
On request
Viral expression
vectors
Mammalian cells
Gene expression in a wide variety of
cell lines, stem cells, non-dividing
cells and tissues
Lentiviral, retroviral and adenoviral systems —
Combination with Tet-regulation, ProteoTuner and fluorescent proteins
On request
BacPaK Baculovirus Expression
System
Insect cells
Baculovirus based gene expression
in insect cells
High protein yield — Proper protein folding —
Expression of biologically active protein with
eukaryotic posttranslational modifications
On request
pAUR vectors
Yeast, filamentous fungi and
bacteria
Expression in yeast
Vectors include a novel drug-resistance selective marker On request
that confers Aureobasidin A resistance to yeast or
filamentous fungal species
pET vectors
Bacteria
Inducible expression of His-tagged
proteins
Strong expression from T7 lac promoter —
N-and C-terminal fusion proteins —
Kits with purification resins included
On request
pCold vectors
Bacteria
Expression of difficult proteins that
cannot be expressed with T7 system
Expression based on cold shock expression
technology — High purity, high yield —
Increased solubility — Vectors with chaperone tags
for high yield of active protein
On request
pNC-His secretory expression
vectors
Brevibacillus
bacteria
Expression of secreted, His-tagged
proteins
Efficient production of secreted or intracellular target
proteins — Brevibacillus produces negligible amounts
of extracellular proteases and no endotoxins —
Proteins are expressed in active form
On request
pRI 101 DNA
vectors
Plants
Stable expression in plants
Expression from cauliflower mosaic virus promoter —
Increased expression by translation enhancer region
On request