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Suitability of genomic DNA synthesized by strand displacement amplification (SDA) for AFLP analysis: genotyping single spores of arbuscular mycorrhizal (AM) fungi.

Abstract：

:Limited biological samples of microbial origin often yield insufficient amounts of genomic DNA, making application of standard techniques of genetic analysis, like amplified fragment length polymorphism (AFLP), virtually impossible. The Phi29 DNA polymerase based whole genome amplification (WGA) method has the potential to alleviate this technical bottleneck. In the present work, we have sought to investigate the suitability of genomic DNA synthesized using Phi29 based WGA for AFLP analysis. We first used genomic DNA from Saccharomyces cerevisiae to optimize the protocol for the use of SDA-amplified DNA for AFLP analysis. Based on the optimized protocol we obtained AFLP fingerprints which were indistinguishable from the non-amplified genomic DNA. Finally, AFLP analysis was performed using SDA synthesized genomic DNA from single spores of various species of arbuscular mycorrhizal (AM) fungi. Unique and highly reproducible fingerprints for each species were obtained. The present study introduces the application of WGA-mediated AFLP to AM fungal biology; similarly, our protocol could be useful for other microbial genomes currently not amenable to genetic analysis owing to the paucity of starting template.

journal_name

J Microbiol Methods

journal_title

Journal of microbiological methods

authors

Gadkar V,Rillig MC

doi

10.1016/j.mimet.2005.03.009

subject

Has Abstract

pub_date

2005-11-01 00:00:00

pages

157-64

issue

2

eissn

0167-7012

issn

1872-8359

pii

S0167-7012(05)00082-5

journal_volume

63

pub_type

杂志文章

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