Rapid polymerase chain reaction (PCR)-single-stranded conformational polymorphism (SSCP) screening method for the identification of Aspergillus section Nigri species by the detection of calmodulin nucleotide variations

Abstract

Single-stranded conformational polymorphism (SSCP) analysis for genetic diversity studies has been widely applied to detect indirectly sequence differences up to a single base in amplified DNA fragments of the same length, representing an alternative to gene sequencing. In this study SSCP analysis was used to detect sequence variations contained in an about 180-bp region of the calmodulin gene in order to identify Aspergillus section Nigri species. The method described shows that fluorescence-based SSCP analysis by capillary electrophoresis is cheaper and faster than direct sequencing, and suitable for computer-assisted analyses allowing discrimination between the Aspergillus species belonging to the Nigri section: A. aculeatus, Aspergillus ‘atypic uniseriate’, A. brasiliensis, A. carbonarius, A. ellipticus, A. foetidus, A. heteromorphus, A. ibericus, A. japonicus, A. niger, and A. tubingensis.

Keywords:

• Aspergillus section Nigri
• single-stranded conformational polymorphism (SSCP)
• species detection
• calmodulin gene
• rapid method

Acknowledgements

This research was supported by the Italian Ministry of Education, University and Research (MIUR) Project No. 12818 — SIVINA — and by European Union Project No. FP6-IST1-508774-IP — GoodFood. The authors thank A. Logrieco for critical suggestions in study coordination.