Analysis of bacterial communities on historical glass by denaturing gradient gel electrophoresis of PCR-amplified gene fragments coding for 16S rRNA

Rölleke, Sabine; Gurtner, Claudia; Drewello, Ursula; Drewello, Rainer; Lubitz, Werner; Weißmann, R.

doi:10.1016/s0167-7012(99)00015-9

Cited by 61 publications

(32 citation statements)

References 19 publications

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“…Thus, sequence information from manually excised DGGE bands, as compared to 1500 bp of the whole 16S DNA sequence, does not always allow reliable phylogenetic analyses. Furthermore, comigration of several different sequences that have the same melting behaviour and therefore the same position in the gel, leads to overlapping DGGE bands, which cannot be sequenced directly (Rö lleke et al, 1999). Alternatively, some studies chose to make a clone library containing fragments of 1500 bp of the 16S ribosomal gene and match clones with specific bands on DGGE gels, with the clones matching with specific bands on DGGE gels further sequenced (Zoetendal et al, 1998;Konstantinov et al, 2003;Zhu et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

DGGE and 16S rDNA analysis reveals a highly diverse and rapidly colonising bacterial community on different substrates in the rumen of goats

Sun

Mao

Yao

et al. 2008

Animal

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In the rumen, plant particles are colonised and degraded by the rumen micro-organisms. Although numerous important findings about fibre-associated bacterial community were obtained using traditional or molecular techniques, little information is available on the dynamics of bacteria associated with feed particles during incubation in the rumen. In the present study, ryegrass leaf, ryegrass stem and rice straw, representing different carbohydrate compositions, were used as substrates and placed in the rumen of goats by using nylon bags, and PCR/DGGE (denaturing gradient gel electrophoresis) with subsequent sequence analysis were used to monitor the dynamics of and identify bacteria associated with the substrates during 24 h of incubation. DGGE results showed that substrate samples collected from 10 min to 6 h had similar DGGE patterns, with up to 24 predominant bands to each sample, including 14 common bands to all samples, suggesting a rapid and stable colonisation by a highly diverse bacterial community. Substrate samples collected at 12 and 24 h showed similar DGGE patterns but had great difference in DGGE patterns from those collected at 10 min to 6 h, suggesting an apparent shift in bacterial community. Sequence analysis indicated that most substrate-associated bacteria were closely related to fibrolytic bacteria. In conclusion, a highly diverse and similar rumen bacterial community could immediately colonise to different substrates and remained stable during the initial 6 h of incubation, but experienced a marked change after 12 h of incubation. Italian ryegrass leaf, Italian ryegrass stem and rice straw were colonised with a similar bacterial community.

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Section: Discussionmentioning

confidence: 99%

DGGE and 16S rDNA analysis reveals a highly diverse and rapidly colonising bacterial community on different substrates in the rumen of goats

Sun

Mao

Yao

et al. 2008

Animal

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“…Along with the development of molecular biology, a number of molecular methods such as random amplified polymorphic DNA (RAPD) and denaturing gradient gel electrophoresis (DGGE) have been applied in microbial diversity (Yang et al 2000;Rö lleke et al 1999). These techniques could characterize the microbial communities more accurately on the molecular scale, but the relationship between the genetic diversity and metabolic diversity and whether the genetic diversity could reflect community-level interactions are uncertain.…”

Section: Discussionmentioning

confidence: 99%

Associated impact of inorganic fertilizers and pesticides on microbial communities in soils

Chen

Zhuang

Xie

et al. 2006

World J Microbiol Biotechnol

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This study was designed to examine the reaction of microbial communities to the associated impact of inorganic fertilizers and pesticides during laboratorial simulation for 90 days. Soils were sampled from fields in three different stations of Chinese Ecosystem Research Network (CERN). The metabolic profiles were characterized by using BIOLOG GN2 microplates. The Shannon Index (H) showed that the metabolic diversity of the three types of soil exhibited similar profiles. PCA and CLUSTER further explored the variance in community structures with the same H. There were obvious changes in the structures of microbial communities during the 90-day duration of the work and the three different types of soil showed similar profiles for most of the period of the experiment.

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“…Some numerically dominant bands from the potato rhizosphere community were also not represented in the temperature gradient gel electrophoresis (TGGE) profiles of any of the BIOLOG wells (14). Bacterial communities from historical window glass were also analyzed by using DGGE of PCR-amplified 16S rDNA fragments (31) and were demonstrated to be much more complex than previously believed.…”

Section: Discussionmentioning

confidence: 99%

Microbial phyllosphere populations are more complex than previously realized

Yang

Crowley

Borneman

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

330

229

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Phyllosphere microbial communities were evaluated on leaves of field-grown plant species by culture-dependent and -independent methods. Denaturing gradient gel electrophoresis (DGGE) with 16S rDNA primers generally indicated that microbial community structures were similar on different individuals of the same plant species, but unique on different plant species. Phyllosphere bacteria were identified from Citrus sinesis (cv. Valencia) by using DGGE analysis followed by cloning and sequencing of the dominant rDNA bands. Of the 17 unique sequences obtained, database queries showed only four strains that had been described previously as phyllosphere bacteria. Five of the 17 sequences had 16S similarities lower than 90% to database entries, suggesting that they represent previously undescribed species. In addition, three fungal species were also identified. Very different 16S rDNA DGGE banding profiles were obtained when replicate cv. Valencia leaf samples were cultured in BIOLOG EcoPlates for 4.5 days. All of these rDNA sequences had 97-100% similarity to those of known phyllosphere bacteria, but only two of them matched those identified by the culture independent DGGE analysis. Like other studied ecosystems, microbial phyllosphere communities therefore are more complex than previously thought, based on conventional culture-based methods.A ll plant species in natural habitats have associated epiphytic microflora comprising the so-called phyllosphere (1, 2). The composition and quantity of nutrients, including carbohydrates, organic acids, and amino acids that support the growth of epiphytic bacteria, are affected by the plant species, leaf age, leaf physiological status, and the presence of tissue damage (3). Similarly, host plants, leaf age, leaf position, physical environmental condition, and availability of immigrant inoculum have also been suggested to be involved in determining species of microbes in the phyllosphere (4-7).There has been much interest in life forms that inhabit extreme environments such as the phyllosphere. With the repeated, rapid alteration of environmental conditions occurring on leaf surfaces, the phyllosphere has been recognized as a hostile environment to bacteria (8). Leaf surfaces are often dry and temperatures can reach 40-55°C under intense sunlight. During the night, however, leaves are frequently wet with dew and at relatively cool temperatures (5-10°C). Strong UV radiation during the day and sparse nutritional (oligotrophic) conditions also contribute to stressful conditions in the phyllosphere (8). More than 85 different species of microorganisms in 37 genera have been reported in the phyllospheres of rye, olive, sugar beet, and wheat, all by culture-based methods (8-10). Most of these bacteria establish large populations with no apparent effect on the plant, but a few of them can infect the leaves and cause disease (1).Microbial ecologists have devoted much effort to investigating microbial diversity and studying biological interactions between species in the environment. Microor...

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Analysis of bacterial communities on historical glass by denaturing gradient gel electrophoresis of PCR-amplified gene fragments coding for 16S rRNA

Cited by 61 publications

References 19 publications

DGGE and 16S rDNA analysis reveals a highly diverse and rapidly colonising bacterial community on different substrates in the rumen of goats

DGGE and 16S rDNA analysis reveals a highly diverse and rapidly colonising bacterial community on different substrates in the rumen of goats

Associated impact of inorganic fertilizers and pesticides on microbial communities in soils

Microbial phyllosphere populations are more complex than previously realized

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