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Identification and Characterization of Microsatellite from Alternaria brassicicola to Assess Cross-Species Transferability and Utility as a Diagnostic Marker.
MedLine Citation:
PMID:  25048820     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
Alternaria blight caused by Alternaria brassicicola (Schwein.) Wiltshire and A. brassicae (Berk.) Sacc., is one of the most important disease of rapeseed-mustard, characterized by the formation of spots on leaves, stem, and siliquae with premature defoliation and stunting of growth. These two species are very difficult to differentiate based on disease symptoms or spore morphology. Therefore, the aim of present investigation was to identify and characterize transferable microsatellite loci from A. brassicicola to A. brassicae for the development of diagnostic marker. A total of 8,457 microsatellites were identified from transcript sequences of A. brassicicola. The average density of microsatellites was one microsatellite per 1.94 kb of transcript sequence screened. The most frequent repeat was tri-nucleotide (74.03 %), whereas penta-nucleotide (1.14 %) was least frequent. Among amino acids, arginine (13.11 %) showed maximum abundance followed by lysine (10.11 %). A total of 32 alleles were obtained across the 31 microsatellite loci for the ten isolates of A. brassicicola. In cross-species amplifications, 5 of the 31 markers amplified the corresponding microsatellite regions in twenty isolates of A. brassicae and showed monomorphic banding pattern. Microsatellite locus ABS28 was highly specific for A. brassicicola, as no amplification was observed from twenty-nine other closely related taxa. Primer set, ABS28F/ABS28R, amplified a specific amplicon of 380 bp from all A. brassicicola isolates. Standard curves were generated for A. brassicicola isolate using SYBR Green I fluorescent dye for detection of amplification in real-time PCR assay. The lowest detection limit of assay was 0.01 ng. Thus, the primer set can be used as diagnostic marker to discriminate and diagnose A. brassicicola from synchronously occurring fungus, A. brassicae associated with rapeseed and mustard.
Authors:
Ruchi Singh; Sudheer Kumar; Prem Lal Kashyap; Alok Kumar Srivastava; Sanjay Mishra; Arun Kumar Sharma
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2014-7-22
Journal Detail:
Title:  Molecular biotechnology     Volume:  -     ISSN:  1559-0305     ISO Abbreviation:  Mol. Biotechnol.     Publication Date:  2014 Jul 
Date Detail:
Created Date:  2014-7-22     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9423533     Medline TA:  Mol Biotechnol     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
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