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Dinoflagellate Bioluminescence: Cloning and Sequence Analysis of Luciferase from Pyrocystis fusiformis
Author(s) -
Ngo Phong D,
Haines Donovan C,
Loeffler Paul A
Publication year - 2011
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.25.1_supplement.lb127
Subject(s) - luciferase , bioluminescence , luciferin , dinoflagellate , biology , cloning (programming) , luciferases , biochemistry , genomic dna , dna , gene , botany , transfection , computer science , programming language
Pyrocystis fusiformis is a protozoan species, approximately forty micrometers in length, that is closely related to plants and algae. These dinoflagellates are photosynthetic and also utilize a luciferase bioluminescence system. Luciferases are a class of enzymes that are responsible for luminescence by catalyzing the oxidation of a substrate molecule, luciferin, in a reaction that produces light at 475 nm. In an effort to discover the structural biology of the luciferase contained within Pyrocystis fusiformis, long‐term research is focused on cloning the genes that code luciferase and purifying luciferin. Alignments of the sequence of this luciferase with other characterized dinoflagellate luciferase allowed the identification of a catalytic core and related fragments for later studies. Extraction and purification of chromosomal DNA from Pyrocystis fusiformis was performed for the purposes of PCR cloning. Dinoflagellates are thecate, meaning that their cellular membranes are interlaced with polysaccharide deposits that form rigid plates making it incredibly difficult to lyse, short of homogenization. Several DNA extraction methods and combinations were tested in order to determine the most effective protocol for preparation of genomic DNA for PCR.