October
2007
, Volume
91
, Number
10
Pages
1,271
-
1,276
Authors
Yong-Yan Chen, Agriculture and Agri-Food Canada (AAFC), Morden Research Station, Unit 100-101, Route 100, Morden, MB, R6M 1Y5; and College of Bioengineering, Dalian University, Dalian 116622, China;
R. L. Conner, AAFC, Morden Research Station, Unit 100-101, Route 100, Morden, MB, R6M 1Y5;
C. L. Gillard, Ridgetown Campus of the University of Guelph, 120 Main St. East, Ridgetown, ON, N0P 2C0;
G. J. Boland, Department of Environmental Biology, University of Guelph, Guelph, ON, N1G 2W1;
C. Babcock, Canadian Collection of Fungal Cultures, AAFC, Room 1015, K.W. Neatby Bldg., Ottawa, ON, K1A OC6;
Kan-Fa Chang, Field Crop Development Center, Alberta Agriculture, Food and Rural Development (AAFRD), Lacombe, AB, T4L 1W1;
S. F. Hwang, Crop Diversification Centre North, AAFRD, Edmonton, AB, T5Y 6H3; and
P. M. Balasubramanian, AAFC, Morden Research Station, Unit 100-101, Route 100, Morden, MB, R6M 1Y5
Affiliations
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RelatedArticle
Accepted for publication 3 May 2007.
Abstract
ABSTRACT
To facilitate early diagnosis and improve control of bean anthracnose, a rapid, specific, and sensitive polymerase chain reaction (PCR)-based method was developed to detect the causal agent, Colletotrichum lindemuthianum, in bean (Phaseolus vulgaris) seed. Based on sequence data of the rDNA region consisting of the 5.8S gene and internal transcribed spacers (ITS) 1 and 2 of four C. lindemuthianum races and 17 Colletotrichum species downloaded from GenBank, five forward primers were designed and evaluated for their specificity. Among them, one forward primer was selected for use in combination with ITS4 to specifically detect C. lindemuthianum. A 461-bp specific band was amplified from the genomic DNA template of 16 representative isolates of C. lindemuthianum, but not from 58 representative isolates of 17 other Colletotrichum species or 10 bean pathogens. Moreover, to enhance the sensitivity of detection, nested PCR was applied, which allowed the detection of as little as 10 fg of C. lindemuthianum genomic DNA and 1% infected seed powder, which was mixed with 99% healthy seed powder. The diagnostic analysis can be completed within 24 h, compared with about 2 weeks required for culturing. Furthermore, this method can be performed and interpreted by personnel with no specialized taxonomic expertise.
JnArticleKeywords
Additional keywords:PCR-based detection
Page Content
ArticleCopyright
The American Phytopathological Society, 2007
