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PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2004-1-27
pubmed:abstractText
A method was developed for genome analysis of phytoplasmas, bacterial plant pathogens that cannot be cultivated in vitro in cell-free media. The procedure includes a CsCl-bisbenzimide gradient buoyant centrifugation followed by polymerase chain reaction (PCR)-mediated whole genome amplification. The latter step involves digestion of the DNA by a restriction enzyme with an A/T-rich recognition sequence. Due to the different A/T content in the DNA of the pathogen and its plant host, the fragments originating from phytoplasma are shorter and are preferentially amplified in the PCR reaction. Products obtained were cloned and screened by dot-blot hybridization. Results showed that about 90% of recombinant clones appeared to harbor phytoplasma specific DNA inserts. Sequencing of randomly selected clones was carried out and comparison with the NCBI database confirmed the bacterial origin for the sequences, which have been assigned a putative function. The origin of the recombinant clones was further confirmed by the generation of specific amplicons from the phytoplasma-infected plant and not from the healthy control, using PCR primers devised from the sequences of the recombinant clones. This method could be used for genome-wide comparisons between phytoplasmas.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Feb
pubmed:issn
0167-7012
pubmed:author
pubmed:issnType
Print
pubmed:volume
56
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
231-42
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
PCR-mediated whole genome amplification of phytoplasmas.
pubmed:affiliation
Departament de Protecció Vegetal, Institut de Recerca i Tecnologi;a Agroalimentaries (IRTA), 08348 Cabrils, Barcelona, Spain. firrao@uniud.it
pubmed:publicationType
Journal Article