Volume 17 Part 1 Article 7: Nucleotide Sequence Polymorphisms of the Partial β-Tubulin Gene in Two Varieties of Pleurotus eryngii

Volume 17 Part 1 Article 7
Year 2008
Title: Nucleotide Sequence Polymorphisms of the Partial β-Tubulin Gene in Two Varieties of Pleurotus eryngii
Authors: A.E. Rodriguez Estrada, D.J. Royse and M.M. Jimenez-Gasco

Abstract:

The β-tubulin gene has been used to reconstruct phylogenetic relationships in fungi at several taxonomical levels. Variation in this gene may allow differentiation between members of the same species based on geographic origin. The purpose of this study was to design and evaluate primers to amplify portions of the β-tubulin gene and to determine nucleotide variation in Pleurotus eryngii var. eryngii and P. eryngii var. ferulae. Two primer pairs used to study phylogenetic relationships in Lentinula spp. and Grifola spp. were evaluated. Additionally, six new primers were designed and used to amplify portions of this gene. The PCR (polymerase chain reaction) amplification that produced a sole, sharp amplicon of 880 bp encompassing three exons and three introns (exon 5 to 8) was selected to amplify and sequence dikaryons of five P. eryngii var. eryngii and three P. eryngii var. ferulae isolates. Nucleotide superimpositions were observed in all P. eryngii var. eryngii and one of the P. eryngii var. ferulae isolates for both coding and non-coding regions. Sequence alignment of all isolates revealed 30 polymorphic sites. In P. eryngii var. ferulae, only one isolate had six sites with nucleotide additivity. However, in P. eryngii var. eryngii, the 30 sites showed additivity in at least one isolate. No indels were detected in the alignment. Two approaches were taken in order to identify the source of polymorphism: sequence analysis of the cloned PCR products and single spore isolates (SSI). Well-defined haplotypes were found to segregate in a Mendelian fashion. Four different alleles were identified in P. eryngii var. eryngii and three in P. eryngii var. ferulae. The results of this study indicate that the β-tubulin gene in P. eryngii is polymorphic and that the alleles are not shared between individuals of the two varieties, suggesting the existence of a reproductive boundary between the groups.

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