ArticleJournal of biomolecular techniques : JBT2007
Optimizing Phytoplasma DNA purification for genome analysis.
Article in Journal of biomolecular techniques : JBT, 2007. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 5 papers.
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Who cites it
5 citing papers in PubMed.
- From vineyard to genome: optimized enrichment and sequencing of Flavescence dorée phytoplasma from grapevine samples.Microbial genomics · 2025Article
- Status and Distribution of Diseases Caused by Phytoplasmas in Africa.Microorganisms · 2025Review
- Iodixanol density gradients as an effective phytoplasma enrichment approach to improve genome sequencing.Frontiers in microbiology · 2022Article
- Multilocus Sequence Analysis Reveals Three Distinct Populations of "Applied and environmental microbiology · 2019Article
- An Effective Pipeline Based on Relative Coverage for the Genome Assembly of Phytoplasmas and Other Fastidious Prokaryotes.Current genomics · 2018Article
Corrections and comments
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Authors and funding
2 authors.
Funding
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Abstract
Genome analysis of uncultivable plant pathogenic phytoplasmas is hindered by the difficulty in obtaining sufficient quantities of phytoplasma enriched DNA. We investigated a combination of conventional enrichment techniques such as cesium chloride (CsCl) buoyant gradient centrifugation, and new methods such as rolling circle amplification (RCA), suppression subtractive hybridization (SSH), and mirror orientation selection (MOS) to obtain DNA with a high phytoplasma:host ratio as the major first step in genome analysis of Candidatus Phytoplasma australiense. The phytoplasma:host ratio was calculated for five different plasmid libraries. Based on sequence data, 90% of clones from CsCl DNA enrichment contained chromosomal phytoplasma DNA, compared to 60% from RCA CsCl DNA and 20% from SSH subtracted libraries. Based on an analysis of representative libraries, none contained plant DNA. A high percentage of clones (80-100%) from SSH libraries contained extrachromosomal DNA (eDNA), and we speculate that eDNA in the original DNA preparation was amplified in subsequent SSH manipulations. Despite the availability of new techniques for nucleic acid amplification, we found that conventional CsCl gradient centrifugation was the best enrichment method for obtaining chromosomal phytoplasma DNA with low host DNA content.
Indexed as
Identifiers
17496222PMC2062537What OpenQuestion holds
Registered trials
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