2012 Promega catalogue

Cell Signaling
Canine Pancreatic Microsomal Membranes
Product Size Cat.# Price ($)
Canine Pancreatic Microsomal Membranes 50 µl Y4041 371.00
For Research Use Only. Not for Use in Diagnostic Procedures.
Description: Microsomal vesicles are used to study co-translational and initial
post-translational processing of proteins. Processing events such as signal
peptide cleavage, membrane insertion, translocation and core glycosylation can
be examined by the translation of the appropriate mRNA in vitro in the presence
of these microsomal membranes. In addition, processing and glycosylation
events may be studied by the transcription/translation of the appropriate DNA in
the TnT ® Lysate Systems when used with Canine Pancreatic Microsomal Membranes.
To assure consistent performance with minimal translational inhibition
and background, microsomes have been isolated free from contaminating
membrane fractions and stripped of endogenous membrane-bound ribosomes
and mRNA. Membrane preparations are assayed for both signal peptidase
and core glycosylation activities using two different control mRNAs. The two
control mRNAs supplied with this system are the precursor for β-lactamase (or
ampicillin resistance gene product) from E. coli and the precursor for α-mating
factor (or α-factor gene product) from S. cerevisiae.
The Signal Sequence Control mRNA (E. coli β-lactamase) is transcribed by SP6
RNA polymerase from a plasmid bearing the coding region for the E. coli gene
encoding the precursor to β-lactamase (the ampicillin resistance gene
product). The RNA is synthesized without a cap analog. This control mRNA
is used to assay for signal peptidase activity and is supplied with the Canine
Pancreatic Microsomal Membranes System.
The Core Glycosylation Control mRNA (S. cerevisiae α-factor) is transcribed by
SP6 RNA polymerase from a plasmid bearing the coding region for the
S. cerevisiae α-mating factor. The RNA is synthesized without a cap analog.
This control mRNA is used to assay for core glycosylation activity and is
supplied with the Canine Pancreatic Microsomal Membranes System.
Features:
• Reliable: Microsomes are stripped of endogenous membrane-bound
ribosomes and mRNA to ensure consistent performance with minimal
translational inhibition and background. Performance tested in rabbit
reticulocyte lysate.
Storage Conditions: Store at –70°C or below. Membranes are stable at
–70°C for 1 year. After thawing, unused portions should be rapidly refrozen
in liquid nitrogen. No detectable loss of activity results after two freeze-thaw
cycles.
Protocol Part#
Canine Pancreatic Microsomal Membranes Technical Manual TM231
µl CMM
precursor
(~31.5kDa)
processed
(~28.9kDa)
µl CMM
processed
(~30.0kDa)
precursor
(~18.6kDa)
For complete and up-to-date product information visit: www.promega.com/catalog
Signal Processing
0 0.3 0.6 0.9 1.2 1.5 1.8 2.1 2.4
Glycosylation
0 0.3 0.6 0.9 1.2 1.5 1.8 2.1 2.4
Processing and glycosylation activity of Canine Pancreatic
Microsomal Membranes (CMM). The positive control mRNAs (0.5µg
each of E. coli β-lactamase and S. cerevisiae α-factor) were translated
using Rabbit Reticulocyte Lysate in a 25μl reaction for 60 minutes in the
presence of the indicated amounts of CMM (3μl). Translation products
were analyzed by gel electrophoresis followed by autoradiography of the
[ 35 S]-labeled proteins.
Amino Acid Mixtures
Product Size Conc. Cat.# Price ($)
Amino Acid Mixture, Complete 175 µl 1 mM L4461 80.00
Amino Acid Mixture Minus Cysteine
Amino Acid Mixture Minus Methionine and
175 µl 1 mM L4471 80.00
Cysteine 175 µl 1 mM L5511 80.00
Amino Acid Mixture Minus Leucine 175 µl 1 mM L9951 80.00
Amino Acid Mixture Minus Methionine
For Laboratory Use.
175 µl 1 mM L9961 80.00
Description: The Amino Acid Mixture, Complete, is an aqueous solution
containing 1mM each of the 20 essential amino acids. This mixture is compatible
for use in the Flexi ® Lysate, TnT ® Lysate and standard Rabbit Reticulocyte
Lysate Systems as well as in the Wheat Germ Extract and E. coli S30 Systems.
Amino Acid Mixtures are also available lacking cysteine, methionine and
cysteine, leucine or methionine.
Storage Conditions: Store at –70°C.
0373TA07_3B
227
13
Protein Expression and Analysis
Section
Contents
Table of
Contents