Single Nucleotide Polymorphism-Based Diagnostic System for Crop-Associated Sclerotinia Species

Abstract

A molecular diagnostic system using single nucleotide polymorphisms (SNPs) was developed to identify four Sclerotinia species: S. sclerotiorum (Lib.) de Bary, S. minor Jagger, S. trifoliorum Erikss., and the undescribed species Sclerotinia species 1. DNAs of samples are hybridized with each of five 15-bp oligonucleotide probes containing an SNP site midsequence unique to each species. For additional verification, hybridizations were performed using diagnostic single nucleotide substitutions at a 17-bp sequence of the calmodulin locus. The accuracy of these procedures was compared to that of a restriction fragment length polymorphism (RFLP) method based on Southern hybridizations of EcoRI-digested genomic DNA probed with the ribosomal DNAcontaining plasmid probe pMF2, previously shown to differentiate S. sclerotiorum, S. minor, and S. trifoliorum. The efficiency of the SNP-based assay as a diagnostic test was evaluated in a blind screening of 48 Sclerotinia isolates from agricultural and wild hosts. One isolate of Botrytis cinerea was used as a negative control. The SNP-based assay accurately identified 96% of Sclerotinia isolates and could be performed faster than RFLP profiling using pMF2. This method shows promise for accurate, high-throughput species identification.

Assessment of Genetic Diversity in Trigonella foenum-graecum Based on Nuclear Ribosomal DNA, Internal Transcribed Spacer and RAPD Analysis

“…The ITS polymorphism might occur at a genus, species or individual levels, making it useful for phylogenetic, evolutionary and bio-geographical diversity studies (Carvalho et al 2009). ITS sequences have been widely used in several species (Hsiao et al 1994;De Bustos and Jouve 2002;Sharma et al 2002;Andrew and Kohn Linda 2009) for phylogenetic inferences.…”

Sclerotinia rot of Zephyranthes candida caused by Sclerotinia sclerotiorum and Sclerotinia minor

“…is distributed globally and can cause white mold, stem rot, or crown rot on many crops and ornamental plants ( Deng et al, 2021 ). The major species of phythopathological interest in the genus sclerotinia include S. sclerotiorum , sclerotinia trifoliorum , and S. minor ( Marion and Myra, 2009 ). In the present study, the pathogens causing sclerotinia rot Z. candida were S. sclerotiorum and S. minor .…”

PRP8 inteins in species of the genus Botrytis and other ascomycetes

“…Cloning and sequencing of PRP8 inteins from Botrytis species and related genera DNA from Botrytis was isolated as described in Staats et al (2005). DNA from Sclerotiniaceae other than Botrytis was isolated as described in Andrew and Kohn (2009). Overlapping amplicons of the full-length PRP8 inteins were cloned using the pGEM-T Easy vector system (Promega) following the manufacturer's instructions.…”