Navigation Links
A Rapid Method for the Purification of Analytical Grade Proteins From Plants Using Preparative SDS-PAGE and Preparative IEF

Contributed by James V. Anderson and Charles L. Guy, The University of Florida, Institute of Food and Agricultural Sciences


Introduction
BiPs (Immunoglobulin Heavy Chain Binding Proteins, also known as GRP78; Glucose Regulated Proteins) are ER-luminal proteins and members of the HSP70 family.1 BiPs are associated with the transport and folding process of proteins involved in the secretory pathway. BiPs and many other HSP70 proteins can easily be partially purified by affinity chromatography with ATP-agarose. However, due to BiPs ability to bind proteins, we have found it associated with other contaminating proteins over a broad range of salt gradients when using ion-exchange chromatography, and over a broad molecular weight range when using size-exclusion chromatography. Here we report a rapid method for the further purification of affinity selected BiP suitable for amino acid sequencing and antibody production.


Methods
Sample Preparation

Native extracts from 3 week old spinach leaf tissue were taken through 40 and 80% ammonium sulfate (AS) precipitation. The resuspended 80% ammonium sulfate pellet was refractionated by an additional 1 M and 3 M ammonium sulfate precipitation. The resulting 3 M pellet was resuspended in 50 mM Tris-HCl (pH 7.5), 1 mM EDTA, and 0.1 mM DTT (Buffer A) and then dialyzed overnight against two changes of buffer A plus 3 mM MgCl2 (Buffer B). The dialyzed sample was centrifuged at 20,000 x g to remove insoluble material. The clarified sample was loaded onto an ATP-agarose affinity column (1.5 x 10 cm bed volume) equilibrated with buffer B at a flow rate of 0.5 ml/min. S
'"/>

Source:


Page: All 1 2 3 4 5

Related biology technology :

1. Simple, Sensitive, and Rapid Detection of FLAG -Tagged Fusion Proteins
2. A Simple and Rapid Technique to Purify High-Quality Midiprep DNA
3. CastAway Precast Gels for Rapid Automated DNA Sequence Analysis
4. Fluorescence-Based Single-Tube Assays to Rapidly Detect Human Gene Mutations
5. HSVision Molecular Beacon Detection Module Rapidly Detects Herpes Simplex Virus DNA
6. Simple and Rapid Isolation of Plant RNA Using the Eppendorf Perfect RNA, Eukaryotic, Maxi Kit
7. Rapid PCR* Using Active Tube Temperature Control On Thermos PCR Machines
8. Molecular Biology:Easy Dephosphorylation and Rapid DNA Ligation
9. Rapid detection of genomic duplications and deletions using the Agilent 2100 bioanalyzer
10. Rapid hit confirmation and early hERG liability testing using the IonWorks HT system
11. Rapidly Prepare Probes and Markers for Small RNA Analysis
Post Your Comments:
(Date:7/10/2014)... Senior supply chain management executives from over ... challenges of “Reducing Cost, Lead Time, & Defects in ... Describing the partnership of the Bio Supply Management Alliance ... ( SCMI) of the University of San Diego ... Kroc Institute of Peace and Justice in San Diego, ...
(Date:7/10/2014)... , July 10, 2014  Kainos Capital, a firm ... today announced that it has acquired the Slim-Fast brand ... the business. Terms of the transaction were not disclosed. ... meal replacement business that markets ready-to-drink shakes, powders, bars ... America and in the United ...
(Date:7/10/2014)... July 10, 2014   Ceres, Inc . (Nasdaq: ... company, announced today financial results for the three months ... its business. Ceres reported that the company ... performance this growing season in Brazil, which concluded in ... company,s sorghum evaluation areas for part of the growing ...
(Date:7/10/2014)... microscopic polymer light resonators that expand in the ... Photonics Laboratory have developed new optical sensors with ... sensors are ideal for detecting trace gas concentrations ... nature, and immunity to electromagnetic interference. , Although ... the MIT team conceived an extremely sensitive, compact ...
Breaking Biology Technology:Biotech & Biopharm Supply Chain Management Executives Launch BSMA in San Diego 2Biotech & Biopharm Supply Chain Management Executives Launch BSMA in San Diego 3Biotech & Biopharm Supply Chain Management Executives Launch BSMA in San Diego 4Kainos Capital Acquires Slim-Fast From Unilever 2Ceres Announces Fiscal Third Quarter 2014 Financial Results 2Ceres Announces Fiscal Third Quarter 2014 Financial Results 3Ceres Announces Fiscal Third Quarter 2014 Financial Results 4Ceres Announces Fiscal Third Quarter 2014 Financial Results 5Ceres Announces Fiscal Third Quarter 2014 Financial Results 6Ceres Announces Fiscal Third Quarter 2014 Financial Results 7Ceres Announces Fiscal Third Quarter 2014 Financial Results 8Ceres Announces Fiscal Third Quarter 2014 Financial Results 9Swell new sensors 2
... ILI ), announced today that it will host ... at 4:30 p.m. ET to review the Company,s,first quarter ... developments., To access the live call, dial 877-627-6555 ... on the Investors,section of the Company,s website, http://www.ilgenetics.com ...
... Pharmaceuticals, Inc.,(Nasdaq: AMLN ) will be presenting at ... 2008 at 10:00 a.m. PT in Las Vegas. Michael,York, ... providing a corporate overview., The live presentation and ... made available following the event. The webcast and,recording will ...
... to augment Quintiles Consulting offerings, RESEARCH ... Corp. today announced that it has signed ... decision-analytics and market research,consulting firm located in ... business., The acquisition of Eidetics will ...
Cached Biology Technology:Interleukin Genetics Announces Conference Call and Webcast to Discuss First Quarter 2008 Business Results 2Quintiles Signs Agreement to Acquire Eidetics 2
(Date:7/10/2014)... overfishing in the English Channel has resulted in the ... left fishermen ,scraping the barrel, for increasing amounts of ... haddock and many other species at the head of ... removed from the area completely., These are some of ... at Plymouth University, in association with international non-profit research ...
(Date:7/10/2014)... at NJIT,s business incubator, received a $1.4 million federal ... use to staunch bleeding during brain surgery. , Surgeons ... the natural bio-polymer solutions in the gel will cohere ... , The gel can shorten an intracranial surgery ... time for the patient,s skull to be open and ...
(Date:7/10/2014)... that Courtney Aldrich, Ph.D., will head the brand-new, ... editor-in-chief. With the first issue slated for publication ... a growing demand for a place to publish ... is to develop a journal focused on diseases ... that advances the field and lays the foundation ...
Breaking Biology News(10 mins):Overfishing in the English Channel leaves fisherman scraping the bottom of the barrel 2Overfishing in the English Channel leaves fisherman scraping the bottom of the barrel 3A start-up at NJIT develops bleeding-control gel for brain surgery 2ACS Infectious Diseases: Unique chemistry journal names editor 2
... from a puddle or a pond to a vast ocean, ... animals. These so-called sessile suspension feeders are critical to aquatic ... contaminants by consuming bacteria. A study published by Cell Press ... that by actively changing the angle of their bodies relative ...
... time, it was thought impossible to isolate a pearl,s ... this elusive goal. Scientists Joana Meyer, from the ETH ... McDonald, and Laurent Cartier of the Swiss Gemmological Institute ... the first time in extracting trace amounts of DNA ...
... Oct. 7, 2013 ValidSoft, a global supplier of ... and a wholly owned subsidiary of Elephant Talk Communications ... commerce technology with Spindle,s new "MeNetwork360" mobile commerce ecosystem, ... ) conference being held from October 6 to October ...
Cached Biology News:How tiny organisms make a big impact on clean water 2A bad break for fake pearls 2ValidSoft and Spindle to Demo New Mobile Commerce Ecosystem at Money2020 2ValidSoft and Spindle to Demo New Mobile Commerce Ecosystem at Money2020 3ValidSoft and Spindle to Demo New Mobile Commerce Ecosystem at Money2020 4ValidSoft and Spindle to Demo New Mobile Commerce Ecosystem at Money2020 5
... TOPO Cloning is ... available for cloning PCR ... and Zero Blunt TOPO ... and activated with topoisomerase ...
ready-to-use solution...
... analysis & visualization software. ... multiple samples. Combine Sequest ... increased confidence. Get more ... from your MS/MS experiments.14 ...
... Immunogen: Synthetic peptide derived from ... rat mouse dog and orangutang occludin ... human occludin protein. Reactivity: Human ... Reactivity with rat mouse dog and ...
Biology Products: