Analysis of the codons for gene expression in rumen protozoa

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This research was conducted to clarify whether rumen protozoa use TGA as a cole stop codon like Tetrahymena living in the natural habitat. Methodologically, genes cloned from cDNA library prepared from a rumen protozoon, Entodinium caudatum. were analyzed and following results were obtained. At first, we succeeded in cloning a chitinase gene (ECchi2). Then the gene was analyzed and the stop condon of the ORF in this gene was revealed to be TAG different from the genes of Tetrahymena. TGA known as a sole stop codon in Tetrahymena was found at the down stream of poly (A) tail in the 3'-terminal region and was thought not to relate with the expression of of the chitinase gene. The gene obtained next time was a tubulin gene (ECtub1). In this ORF.TAA was recognized as a stop codon through the universal codon and TGA located at the down streem of the TAA.However, there existed CAA which recognized glutamine (Q) just before the TAA and furthermore there were six more CAA codons for glutamine. Thus TAA was thought to be used as a stop codon in this ORF.It was also considered that the tubulin gene has a high possiblity to use universal codons recognizing TAA as a stop codon from the example of chitinase. A centrin gene which is a gene of another constituents of microtubule in the cells together with tubulin was already been cloned though the joint research project between Miyazaki and UK and it was revealed that a stop codon in this gene was TAA.The gene cloned finally was acid phosphatase gene (ECaph1) and a stop codon of this ORF was TAA.In this gene, ten glutamine was included and nine of them were expressed by CAA and another one by CAG.Thus TAA was thought to be used as stop codon. From all these results, rumen protozoon, Entodinium caudatum, was highly considered to use universal codons.