Mehmann B, Brunner I, Braus G H
Institute of Microbiology, Swiss Federal Institute of Technology, Zürich.
Appl Environ Microbiol. 1994 Sep;60(9):3105-11. doi: 10.1128/aem.60.9.3105-3111.1994.
DNA sequences of single-copy genes coding for chitin synthases (UDP-N-acetyl-D-glucosamine:chitin 4-beta-N-acetylglucosaminyltransferase; EC 2.4.1.16) were used to characterize ectomycorrhizal fungi. Degenerate primers deduced from short, completely conserved amino acid stretches flanking a region of about 200 amino acids of zymogenic chitin synthases allowed the amplification of DNA fragments of several members of this gene family. Different DNA band patterns were obtained from basidiomycetes because of variation in the number and length of amplified fragments. Cloning and sequencing of the most prominent DNA fragments revealed that these differences were due to various introns at conserved positions. The presence of introns in basidiomycetous fungi therefore has a potential use in identification of genera by analyzing PCR-generated DNA fragment patterns. Analyses of the nucleotide sequences of cloned fragments revealed variations in nucleotide sequences from 4 to 45%. By comparison of the deduced amino acid sequences, the majority of the DNA fragments were identified as members of genes for chitin synthase class II. The deduced amino acid sequences from species of the same genus differed only in one amino acid residue, whereas identity between the amino acid sequences of ascomycetous and basidiomycetous fungi within the same taxonomic class was found to be approximately 43 to 66%. Phylogenetic analysis of the amino acid sequence of class II chitin synthase-encoding gene fragments by using parsimony confirmed the current taxonomic groupings. In addition, our data revealed a fourth class of putative zymogenic chitin synthesis.
编码几丁质合成酶（UDP-N-乙酰-D-葡糖胺：几丁质4-β-N-乙酰葡糖胺基转移酶；EC 2.4.1.16）的单拷贝基因的DNA序列被用于表征外生菌根真菌。从约200个氨基酸的产酶几丁质合成酶区域两侧的短的、完全保守的氨基酸片段推导而来的简并引物，使得该基因家族的几个成员的DNA片段得以扩增。由于扩增片段的数量和长度存在差异，担子菌纲真菌获得了不同的DNA条带模式。对最突出的DNA片段进行克隆和测序后发现，这些差异是由于保守位置存在各种内含子。因此，担子菌纲真菌中内含子的存在有可能通过分析PCR产生的DNA片段模式来用于属的鉴定。对克隆片段的核苷酸序列分析显示，核苷酸序列的差异在4%到45%之间。通过比较推导的氨基酸序列，大多数DNA片段被鉴定为II类几丁质合成酶基因的成员。同一属物种的推导氨基酸序列仅在一个氨基酸残基上有所不同，而同一分类类别的子囊菌纲和担子菌纲真菌氨基酸序列之间的同一性约为43%至66%。使用简约法对II类几丁质合成酶编码基因片段的氨基酸序列进行系统发育分析，证实了当前的分类分组。此外，我们的数据揭示了第四类假定的产酶几丁质合成。
