Advanced
Analytical Techniques - Oligoprobe Hybridization
Oligoprobe hybridization
is a technique that makes it possible to confirm the genetic identity of microorganisms isolated from mixed populations within the environment. RNA or DNA extracted from these microbes is fixed to a charged membrane such as nitrocellulose or nylon and supplemented with enzyme-labeled probes. Following a short incubation and reaction with specific substrates, the labeling profiles are examined to determine if target species are present.
Oligoprobe
Hybridization for Bioremediation
Chlorinated solvents such as tetrachloroethene & trichloroethene are common to a number of industries and are extremely resistant to degradation in the natural environment. These compounds are being identified increasingly in the subsurface environments near industrial and military installations and may be important groundwater contaminants. Although few microorganisms are able to metabolize these solvents, the dehalococcoides (DHC) group is capable of breaking them down to nontoxic endproducts such as ethenes. However, the ecology and distribution of dehalococcoides in the subsurface is not well understood. Because of the strong interest in promoting the growth of DHC within contaminated soils, SMI has partnered
with firms that specialize in the production and use of DHC for bioremediation applications. SMI scientists have adapted oligoprobe hybridization for use in identifying and quantifying DHC bacteria collected from subsurface samples.
Enumeration of DHC Bacteria Using Oligoprobe Hybridization
A brief description
of the oligoprobe hybridization technique developed by SMI is presented below.
STEP 1:
A water sample is incubated overnight
and colonies are grown on an agar plate.
STEP
2:
A positively charged membrane is applied to the surface of the culture plate. The membrane binds
DNA extracted from the bacterial colonies.
STEP
3:
Acid is added to release the DNA
STEP
4:
Acid is neutralized
STEP
5:
Hybridization - A tag (oligoprobe) is
added and binds
to specific DNA sequences.
STEP
6:
Substrates are added for detection of
desired
colonies (dehalococcoides, DHC)
STEP
7:
Identification / Quantification of Dehalococcoides (DHC) from the heterotrophic population.
SMI
is committed to using innovative technologies to solve environmental
problems. In keeping with this philosophy, SMI is
actively investigating alternative uses for this technology.
For further information about SMI's services, please feel to contact a project manager at (574) 277-4078.
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