By Robert A. LaRossa
A state of the art number of leading edge tools for using luciferase and eco-friendly fluorescent protein in analytical biochemistry, molecular toxicology, environmental tracking, and cell-based assays. The contributors-all hands-on researchers who've perfected their protocols to a excessive measure of reproducibility-illustrate those robust equipment with a large choice of functions starting from the analytical selection (titer) of particular molecules to the mobile detection of a variety of pollution, to microbiological detection and research, and the molecular organic manipulation of lux, luc, and gfp structural genes. step by step equipment also are incorporated for the development of alternative recombinant organisms.
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Extra info for Bioluminescence Methods and Protocols
A. (1994) A yeast RNA-binding protein shuttles between the nucleus and the cytoplasm. Mol. Cell. Biol. 14, 8399-8407. 10. Olson, K. , McIntosh, J. , and Olmsted, J. B. (1995) Analysis of MAP4 function in hving cells using green fluorescent protein (GFP) chimeras. J CeZI. Bzol. 130,639-650 Green Fluorescent Protein Gene 41 11. Kaether, C. -H. (1995) Visualization of protein transport along the secretory pathway using green fluorescent protein. FEBS Lett. 369,267-27 I. 12. , Moss, G. W. , and Hughes, T.
2 (see Note 9). Ford and Leach 62 100000 10000 1000 100 10 Nucleotide, pmoihample Fig. 2 Standard curve for condltlons that permit measurement of 0 1 pmol of guanylate. The procedure described in the text was apphed to standards containing varying concentrationsof ATP, GTP, andGMP. The averagesof duplicatedeterrmnationswere plotted Modified from ref. 23. H ATP, 0 GTP, and l GMP. 8. Guanylafe Standard Curve To determine the amounts of guanylates, a standard curve must be generated to correlate the light production with amount of nucleotide.
5 [GDP] [GTP] + [GDP] + [GMP] (8) Standard samples of known energy charge should be assayedto calibrate the procedure and assurethat there is good agreement between experimental values for standards and theoretical values. 00. 50 “Standard amounts of nucleottdes (10 pmol) were used as Indicated m the first column The counts were obtamed from trtphcate samples treated through steps l-4 as described m the text Columns A, B, and C present m A, GTP; m B, GTP + GDP, and m C, GTP + GDP + GMP GEC was calculated as indicated m the text From ref.
Bioluminescence Methods and Protocols by Robert A. LaRossa