caa a22 4500 467929793 CHVBK 20180406152940.0 cr unu---uuuuu 170328e20060801xx s 000 0 eng 10.1007/s10658-006-9029-0 doi (NATIONALLICENCE)springer-10.1007/s10658-006-9029-0 Sensitive Detection of a Phytoplasma Associated with Little Leaf Symptoms in Withania somnifera [Elektronische Daten] [J. Khan, P. Srivastava, S. Singh] Withania somnifera is an important medicinal plant native to the Indian-sub continent. Owing to the presence of a number of precious alkaloids, flavonoids and withanolides, it is widely used in the Indian and African systems of medicines. It is severely affected by phytoplasma present in the sieve tubes of phloem. With a view to micropropagate phytoplasma-free W. somnifera plants, an efficient and effective nested PCR-based system was developed for detection of associated phytoplasmas. Universal primers, designed from the 16S rDNA sequences of phytoplasmas, were applied in direct/nested-PCR. Total DNA extracts from leaf tissues of 33 suspected symptomatic and 11 non-symptomatic plants were subjected to direct PCR. The direct PCR products were subsequently employed as templates in nested PCR. The nested PCR could reamplify direct PCR products yielding a DNA fragment of 1.4kb. A phytoplasma was detected in all the diseased plants and not from the healthy looking plants. Further, it was sensitive enough to amplify phytoplasma DNA obtained from crude DNA diluted up to 2500 times from naturally infected plants and also from various stages of in vitro-propagated diseased plants. Identical restriction fragment polymorphism enzyme profiles were obtained following restriction enzyme digestion of nested PCR products, obtained from five different plants, by EcoRI, AluI and RsaI restriction endonucleases. The developed nested PCR based system should facilitate indexing of the phytoplasma in different stages of in vitro-generated plants and probably identification of, as yet unknown, hosts and vectors of phytoplasma associated with phytoplasma disease of W. somnifera. Springer, 2006 detection nationallicence elimination nationallicence phytoplasma nationallicence Withania somnifera nationallicence PCR : polymerase chain reaction nationallicence RFLP : restriction fragment length polymorphism nationallicence Khan J. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut Srivastava P. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut Singh S. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut European Journal of Plant Pathology Kluwer Academic Publishers 115/4(2006-08-01), 401-408 0929-1873 115:4<401 2006 115 10658 https://doi.org/10.1007/s10658-006-9029-0 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10658-006-9029-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Khan J. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut NATIONALLICENCE 700 1- Srivastava P. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut NATIONALLICENCE 700 1- Singh S. Plant Virus Laboratory, National Botanical Research Institute, 226 001, Lucknow, India aut NATIONALLICENCE 773 0- European Journal of Plant Pathology Kluwer Academic Publishers 115/4(2006-08-01), 401-408 0929-1873 115:4<401 2006 115 10658 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer