رديابي آلودگي هاي مخلوط فايتوپلاسمايي در بوته هاي سيب زميني

چكيده انگليسي :

94 Detection of Mixed Phytoplasma Infection on Potato Plants Javad Gholami J gholami417@gmail com July 27 2015 Department of Plant Protection Isfahan University of Technology Isfahan 84156 83111 Iran Degree M Sc Language Farsi Supervisor M Bahar mbahar@cc iut ac irAbstractPotato associated phytoplasmas significantly reduce quality and quantity of potato worldwide Among the potatophytoplasmas caused diseases potato purple top Candidatus Phytoplasma asteris and potato witches broom Ca p trifolii with major economic effects are the most prevalent in the world Of the Phytoplasmas recorded inIsfahan Ca P solani with high incidence and both Ca P trifolii and Ca P asteries with low incidence ofoccurrence have been reported from potato fields however no definite correlation was found between diseasedplants symptoms and phytoplasma species This finding suggests the possibility of multi phytoplasmal associationwith each symptomatic plant which may enhance disease severity and yield loss To test the presence of more thanone phytoplasma in individual potato plants exhibiting purple top or whiches broom symptoms The potato plantsshowing symptoms including yellowing stunning purpling top leaf rolling and having aerial tuber and swollenbud were collected from Isfahan Damaneh Daran Fereydun shahr Chadehan and Boin miandasht andChaharmahale Bakhtiari Sharekord and Faradonbeh potato growing area The DNA extracted from leaf midribsand petioles of the samples were tested of by nested PCR using universal primer pair P1 P7 in the first roundfollowed by the second round amplification using either of the specific primer pair R16F2n R2 The PCRamplicons 1239 bp were subjected to RFLP analysis which allowed determining different phytoplasma groupsbased on CfoI digestion pattern Sequence analysis of PCR products of primer pair R16f2n R2 of therepresentative phytoplasmas from each subgroup confirmed the presence of Ca P solani and Ca P trifolii insymptomatic potatoes although Ca P asteris was not associated with the samples studied To develop amultiplex PCR assay for simultaneous detection of mixed phytoplasma infection in diseased plants specific primerpairs were designed based on 16 23S rRNA and SecY gene regions of Ca P solani Ca P trifolii and Ca P asteris Among the primer sets examined in PCR reaction with known phytoplasmas Ast F Ast R 1419 bp Ast F Ast R2 1544 bp and Ast F Ast R3 1450 bp were effective for individual detection of Ca P asteris Primerpairs Ast F m2Sol R 1274 bp and Ast F Sol R3 1410 bp and primer sets TRF1 F TRF R 1379 bp TRF1 F TRF R2 1530 bp and TRF1 F TRF R3 1430 bp were also appropriate for detection of Ca P solani and Ca P trifolii respectively The results of multiplex PCR reaction using Com F m2Sol R TRF R3 Ast R2 primercombination ensured simultaneous detection of three mentioned phytoplasmas by amplification of the expectedbands Of the positive samples tested 30 samples were found infected by Ca P solani two samples wereidentified in association with Ca P trifolii and 15 samples were distinguished as mix infected by both Ca P solani and Ca P trifolii however none of the samples was found contaminated with Ca P asteris Because of thesimilarity between the symptoms of Phytoplasmal and Zebra chip Candidatus Liberibacter solanacearum disease to test the role of Ca Liberibacter solanacearum for development of purple top symptoms the collected sampleswere assessed in nested PCR assay initially using OA2 OI2c primer pair followed by Lib16S01F R However theprimers did not amplify the expected DNA fragments of Zebra chip causal agents in the samples In greenhouseassessment the tubers of phytoplasma associated plants were grown and subjected to multiplex PCR assay forsimultaneous detection of phytoplamas transmission to next generation Of the tubers of 30 infected plants onlyfive samples were found infected with phytoplasma which demonstrates the very low rate of phytoplasmatransmission from plant to tuber and from tubers to next ge

غلامي، جواد

رديابي آلودگي هاي مخلوط فايتوپلاسمايي در بوته هاي سيب زميني

طراحي آغازگر , رديابي توام