Application of random amplified polymorphic DNA (RAPD) analysis coupled with microchip electrophoresis for high-resolution identification of Monascus strains.
Monascus fungi are commonly used for a variety of food products in Asia, and are also known to produce some biologically active compounds. Since the use of Monascus is expected to increase in food industries, strain-level identification and management of Monascus will be needed in the near future. In the present study, random amplified polymorphic DNA (RAPD) analysis coupled with microchip electrophoresis was applied for this purpose. Evaluations of the analysis stability revealed that reproducible results could be obtained, although template DNA fragmentation could influence the resulting RAPD pattern. RAPD analysis using 15 Monascus strains consisting of four species, M. ruber, M. pilosus, M. purpureus, and M. kaoliang showed that each strain generated a unique RAPD pattern, which allows strain-level identification of Monascus. In addition, the phylogenetic tree constructed from RAPD patterns reflected M. ruber-M. pilosus and M. purpureus-M. kaoliang clusters inferred from both ITS and beta-tubulin gene sequences, which indicated that the RAPD pattern could reflect their phylogenetic traits to a certain extent. On the other hand, RAPD analysis did not support the monophyletic clustering of the four Monascus species used in this study, which suggests the necessity of reexamination of species boundaries in Monascus.
Shinzato
Naoya
N
Center of Molecular Biosciences, University of the Ryukyus, 1 Senbaru, Nishihara-cho, Okinawa 903-0213, Japan.
Namihira
Tomoyuki
T
Tamaki
Yasutomo
Y
Tsukahara
Masatoshi
M
Matsui
Toru
T
D016000:Cluster Analysis; D016172:DNA Fingerprinting / Q000379:methods; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000737:chemistry / Q000235:genetics; D046209:Electrophoresis, Microchip / Q000379:methods*; D005838:Genotype; D008969:Molecular Sequence Data; D032741:Monascus / Q000145:classification* / Q000235:genetics*; D016533:Mycological Typing Techniques / Q000379:methods*; D010802:Phylogeny; D019105:Random Amplified Polymorphic DNA Technique / Q000379:methods*; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA; D014404:Tubulin / Q000235:genetics
D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer; D014404:Tubulin