PCR cloning, DNA sequencing and phylogenetic analysis of a xylanase gene from the phytopathogenic fungus Ascochyta pisi Lib

Peter Stephensen Lübeck, Lars Paulin, Yeshitila Degefu, Mette Lübeck, Irina A. Alekhina, Sergey A. Bulat, David B. Collinge

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

A gene encoding a xylanase, xyl1, was isolated from the phytopathogenic fungus Ascochyta pisi Lib. by PCR cloning using degenerate primers. DNA sequence analysis revealed an open reading frame of 736 bp interrupted by an intron of 55 bp. The ORF encodes a predicted protein of 227 amino acids. The precise splicing site of the intron was identified from the sequence of a PCR product obtained using the same degenerated primers on a cDNA template. The cDNA product and a northern blot demonstrated that the gene is transcribed into mRNA when the fungus is cultured in media containing xylan as sole carbon source. The Neighbour - Joining method using the Clustal W(1.5) program demonstrated that the A. pisi xylanase is a member of the family 11 glycosyl hydrolases, and that this family represents at least five phylogenetically consistent groups. The family 11 glycosyl hydrolases can be linked with family 10 glycosyl hydrolyses through bifunctional enzymes from Ruminococcus flavefaciens and, to a lesser extent Neocallimastix patriciarum. (C) 1997 Academic Press Limited.
Original languageEnglish
JournalPhysiological and Molecular Plant Pathology
Volume51
Issue number6
Pages (from-to)377-389
ISSN0885-5765
DOIs
Publication statusPublished - 1997
Externally publishedYes

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