|Iversen, William - Bill|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 12/1/2006
Publication Date: 2/28/2006
Citation: Lartey, R.T., Caesar, T., Hanson, S.L., Iversen, W.M., Evans, R.G. 2006. Detection of Cercospora beticola by PCR in Amended and Naturally Infested Field Soil. American Society of Sugar Beet Technologists, February 28-March 3, 2007, Salt Lake City, Utah. p. 37. Interpretive Summary:
Technical Abstract: The causal agent of Cercospora Leaf Spot of sugar beet (Beta vulgaris L), Cercospora beticola. Sacc. survives as stromata in beet leaf residues in the soil. Under optimal conditions, overwintering propagules germinate and produce conidia that are dispersed as primary inoculum to initiate infection in sugar beet. We developed and present here a PCR technique for detection of C. beticola in the soil. The DNA was purified from soil amended with C. beticola and naturally infested soil using PowerSoil DNA Kit (MO BIO, CA) as per manufactures instructions. The purified DNA was collected and subjected to PCR reaction in Extract-N-Amp PCR mix (Sigma Aldrich, St Louis MO) with CBACTIN and ITS based primers. Amplification was carried out over 35 cycles using a Mastercycler gradient thermocycler (Eppendorf Scientific Inc., Westbury, NY) at 94°C for 1 min denaturation, 52°C for 30 sec annealing, 72°C for 1 min extension and 5 min final extension at 72°C. The amplified products were resolved by electrophoresis in 1% agarose gels. The fragment sizes of C. beticola amended and the infected field soil products correlated with the expected size of the control DNA extracts from C. beticola cultures. Amplicons were sequenced and compared to C. beticola actin sequence from gene bank. Alignment of sequences of the amplified products confirmed them to be that of C. beticola. The system will enable rapid post planting screening for inoculum potential of C. beticola in soil and determine effect of soil applied biocontrol agents on C. beticola and inoculum potential.