Marker Gene Monthly Newsletter   

May, 2004

Volume 4, Number 5

© Copyright MGT, Inc., 2007.  Published by Marker Gene Technologies, Inc., The University of Oregon Riverfront Research Park, 1850 Millrace Drive, Eugene, Oregon 97403-1992 USA.  All rights reserved.  For information on the use or copying of the material contained in this document, please contact us at techservice@markergene.com.  Please see below for subscription information and updates.  This newsletter is labeled as an ADVERTISEMENT in accordance with the CAN-SPAM act of 2003, S.877 Public Law: 108-187.

New Mammalian GFP Vectors.

Researchers at the Institute of Bioorganic Chemistry, Russian Academy of Sciences in Moscow, Russia, have developed a photoactivatable GFP-like protein, generated by site-directed and random mutagenesis of the asulCP chromoprotein from the sea anemone species Anemonia sulcata, that switches from nonfluorescent to a bright red fluorescent form and back in response to light of certain intensity and wavelength.  The protein is photoactivated under green light (530-560 nm) and relaxes to the initial non-fluorescent form (half-life 50 seconds), or can be quenched instantly by blue light (430-490 nm). Many variations of the light response, duration, as well as stable fluorescence versions have been developed.   For more information about these and other new mammalian GFP-like fluorescent protein expression vectors, see the references below or visit our website or the website www.evrogen.com.

• Chudakov D.M., Lukyanov K.A., “Use of green fluorescent protein (GFP) and its homologs for in vivo protein motility studies.” Biochemistry (2003) 68(9): 952-7
• Chudakov D.M., Belousov V.V.,  Zaraisky A.G., Novoselov V.V., Staroverov D.B., Zorov D.B., Lukyanov S., Lukyanov K.A., “Kindling Fluorescent Proteins for precise in vivo photolabeling.” Nature Biotechnology, (2003) 21(2): 191-194.
•  Chudakov D.M., Feofanov A.V., Mudrik N.N., Lukyanov S., Lukyanov K.A.,  “Chromophore environment provides clue to "kindling fluorescent protein" riddle.” Journal of Biological Chemistry, Feb; 278(9): 7215-7219.
•   Lukyanov K.A., Fradkov A.F., Gurskaya N.G., Matz M.V., Labas Y.A., Savitsky A.P., Markelov M.L., Zaraisky A.G., Zhao X., Fang Y., Tan W., Lukyanov S.A.,  “Natural animal coloration can be determined by a nonfluorescent green fluorescent protein homolog.” Journal of Biological Chemistry, 2000, 275(34): 25879-25882.

Nanotechnology’s Quantum Dots Now Appearing in Live Cell Applications.

Quantum dots are nano-sized crystals that exhibit fluorescence emission due to their unique semiconductor qualities.  By altering their size, they can be made to produce narrow bandwidth, multiple wavelengths of emission, using a single wavelength of excitation light. Thus, spectral overlap does not limit the number of colors that can be used at once in an experiment.  In addition, quantum dots (QDs) are quite photostable, and compared to most fluorescent dyes, shine for an average of 1,000 times longer.  But because of their large size (typically 50 nm or more) and hydrophobicity, until recently, quantum dots have only found use in extracellular labeling experiments (antibody, cell surface receptor labeling, etc.), or by microinjection. Several recent reports however, indicate that quantum dots can be internalized into living cells using specific coatings and endocytotic mechanisms.  Endocytosis is a process used by many cell types to internalize larger proteins or nutrients.  Although the internalized QD’s were confined to endocytotic vesciles once inside the cells, these methods present a step forward toward the use of these exciting new fluorescent labels for intracellular detection.  For more information about quantum dots and new labeling applications, visit our website or see the references below:

• Osaki F., Kanamori T., Sando S., Sera T., Aoyama Y., (2004) “A quantum dot conjugated sugar ball and its cellular uptake. On the size effects of endocytosis in the subviral region.” J Am Chem Soc 126(21): 6520-1.
• Jaiswal JK ; Mattoussi H ; Mauro JM ; Simon SM., “Long-term multiple color imaging of live cells using quantum dot bioconjugates.” (2003) Nat Biotechnol  21(1): 47-51.
• Pellegrino T., Parak W.J., Boudreau R., Le Gros M.A., Gerion D., Alivisatos A.P., Larabell C.A., (2003) “Quantum dot-based cell motility assay.” Differentiation 71(9-10): 542-8.

New Aza-Epoxide Peptidase Inhibitors for Apoptosis Research. 

Cysteine proteases, including the Caspases, are involved in numerous important physiological processes in the cell, including cytokine maturation, apoptosis, neurodegeneration and blood coaglulation.  These enzymes employ a nucleophilic thiol in their reactive site, and can thus be targeted by electrophilic groups on specific C-terminal residues of peptides, as suicide inhibitors.  A series of such aza-epoxide derivatives have been synthesized in the laboratories of Dr. Jim Powers at the Parker H. Petit Institute of Bioengineering and Bioscience, Georgia Institute of Technology in Atlanta, GA.   Similar to the fluoromethylketone (FMK) inhibitors, these new inhibitors show improved specificity, distinct enantioselectivity and excellent activity.  The potential uses of these new inhibitors as therapeutic agents and for a variety of disease states are substantial.  Marker Gene is working together with researchers at GIT and Immunochemistry Technologies, Inc. to help develop these new inhibitors for additional research applications.  For more information about these technologies, please see the references below:

• "Aza-peptide Michael acceptors: A new class of inhibitors specific for caspases and other clan CD cysteine proteases," O. D. Ekici, M. G. Götz, K. E. James, Z. Z. Li, B. J. Rukamp, J. L. Asgian, C. R. Caffrey, E. Hansell, J. Dvorák, J. H. McKerrow, J. Potempa, J. Travis, J. Mikolajczyk, G. S. Salvesen, J. C. Powers, J. Med. Chem., 2004, 47.
• "Design, synthesis, and evaluation of aza-peptide epoxides as selective and potent inhibitors of caspases-1, -3, -6, and -8," K. E. James, J. L. Asgian, Z. Z. Li, O. D. Ekici, J. R. Rubin, J. Mikolajczyk, G. S. Salvesen, J. C. Powers, J. Med. Chem., 2004, 47, 1553-1574.
• "Aza-peptide epoxides: Potent and selective inhibitors of schistosoma mansoni and pig kidney legumains (asparaginyl endopeptidases)," K. E. James, M. G. Götz, C. R. Caffrey, E. Hansell, W. Carter, A. J. Barrett, J. H. McKerrow, J. C. Powers, Biol. Chem., 2003, 384, 1613-1618.

New in vivo GUS Tissue Assays

Marker Gene, in collaboration with researchers at The University of Oregon and the USDA-ARS-Northern Crop Science Laboratory are developing a new tissue assay system for analysis of the GUS reporter gene in plant leaf disk samples.  The assay is quick and sensitive, and provides the ability to directly analyze leaf disk samples soaked in a perfusion buffer for GUS activity.  The proprietary enhanced-permeability substrate used in the kit, improves plant cell penetration.  The assay system is based upon a microtiterplate format, and can be automated for routine analyses.  Please see our web site below for information on this new assay system (available soon) or our other GUS marker gene assay products (M0240, M0877).

• Jefferson R.A., Burgess S.M., Hirsh D.,  (1986) “beta-Glucuronidase from Escherichia coli as a gene-fusion marker.” Proc. Natl. Acad. Sci. USA. 83(22): 8447-51.
• Jefferson R.A., Kavanagh T.A., Bevan M.W., (1987)  “GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants.” EMBO J. 6: 3901-3907.
• Kosugi S., Suzuka I., Ohashi Y., Murakami T., Arai Y.,  (1991) “Upstream sequences of rice proliferating cell nuclear antigen (PCNA) gene mediate expression of PCNA-GUS chimeric gene in meristems of transgenic tobacco plants.” Nucleic Acids Res 19(7): 1571-6.
• Gallagher, S.R.,ed. (1992) “GUS Protocols: Using the GUS Gene as a Reporter of Gene Expression”. Academic Press, New York, NY.
  
  

Waveguide Fluorescent Assay Systems.

Total Internal Reflection Fluorescence (TIRF) is a relatively new technique that utilizes the optical properties of glass (microscope slides) and other media to selectively excite fluorescent molecules that are inside extremely thin optical sections of living cells.  By focusing the exciting light at a critical angle (inside the glass slide or cover slip on which the cells are attached) an evanescent wave is created that only penetrates adherent cells in the thin section (about 100 nm) in contact with the coverglass.  The evanescent wave typically only reaches this short distance into the specimen and then its energy drops off exponentially. Because the specimen is not excited beyond the evanescent wave, this imaging system can produce fluorescence images with an extremely high signal-to-noise (S/N) ratio.  TIRF systems have also been used for adherent antibody assays, proteomic and genomic microarray “chip” assays.  For more information about these exciting new analytical techniques, see the references below:

• Davidson, M. W., Axelrod, D., Florida State Univ. server at http://micro.magnet.fsu.edu/primer/techniques/fluorescence/tirf/tirfconfiguration.html
• Ruckstuhl T., Seeger S., “ Attoliter detection volumes by confocal total-internal-reflection fluorescence microscopy.” Opt. Lett. (2004) 29(6): 569-71.
  
  

2004-2005 Catalog Will Be Available Soon.

The 2004-2005 edition of the Marker Gene catalog is in production.  Many new products and kits, additional literature references, data and protocols will be included, as well as new information about our old products.  Be sure to add your name to our mailing list.  Please visit our Web site and fill out our Customer Information Form, or e-mail us at techservice@markergene.com and we will have a copy sent out to you.
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Compare Our Quality. 

Marker Gene strives to offer our customers products of the highest quality and at the best possible prices.  Our years of experience allow us to provide timely products for less cost to you.  See our latest Price Comparison Chart that compares our prices with those from several alternate sources, to see if you can save money by switching to Marker Gene (http://www.markergene.com/crossref.htm).  Or visit our website at www.markergene.com and click on the link “COMPARE”.  We think you will appreciate our efforts to keep costs low and maintain excellent quality of our products for your research.  For more information about any of our products, simply telephone us toll free at 1-888-218-4062 or contact us by e-mail at techservice@markergene.com.  We will be happy to send you more about our products and their specifications.

CONTRACT  RESEARCH@markergene.com

Marker Gene Technologies, Inc. has the expertise to perform contract research with you on your project. We have worked with many biotechnology and pharmaceutical companies on successful, proprietary and patented projects.

Contract Research and Development Capabilities in the following areas:

• Established in 1993 at the University of Oregon Riverfront Research Park.
• Screening Assay Development for HTS and uHTS
• Chemical and Cellular Assays – High-Content Screening.
• DNA/RNA (genomics) and protein (proteomics) labeling and assay development.
• Pharmaceutical Intermediates - design, synthesis, and in vitro testing in mammalian cell culture.
• Specializing in Carbohydrate, Lipid, Peptide, and Nucleic Acid Chemistries.
• Fully equipped laboratories (Biochemistry, Chemical Synthesis, Tissue Culture, Analytical).
• Confidentiality, help in patent preparation and filings.

Contact us by telephone at (888) 218-4062 or (541) 342-3760 or FAX us at (541) 342-1960 or you can write to us at  Contract Research, Marker Gene Technologies, Inc., 1850 Millrace Drive, Eugene, Oregon 97403-1992 or contact us by e-mail at: techservice@markergene.com

Marker Gene Accepts Major Credit Cards.

Place your orders now, using Master Card or Visa and save time and money!  Our Customer Assistance Staff can now accept either Master Card or Visa Credit Card orders, securely by telephone (toll-free) at 1-888-218-4062 (Domestic orders only).   We will continue to accept Institutional Purchase Orders for our products, online or by FAX at 1-541-342-1960.  International customers should contact us by e-mail, post or telephone for more information about International Distributors and ordering.  For information on pricing for individual products, or for a quote on bulk quantities of our products or kits, please contact our technical assistance staff at techservice@markergene.com.   We will be happy to assist you. 

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