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BISC209: Enrichment: Difference between revisions
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→Enrichment media and Isolation protocols (Starting with SOIL)
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<UL><LI> | <UL><LI> | ||
'''Enrichment:'''<BR> | '''Enrichment:'''<BR> | ||
Measure 0.5 g of soil (collected in LAB 2) into 25 ml of liquid | Measure 0.5 g of soil (collected in LAB 2) into 25 ml of liquid Azotobacteria medium. (This is already aliquoted for you in small flasks with cotton plugs or a loose cover.) <LI> | ||
Mix well<LI> | Mix well<LI> | ||
Place the flask in your closed bench cabinet so the culture will incubate in the dark at RT. </LI></UL> | Place the flask in your closed bench cabinet so the culture will incubate in the dark at RT. </LI></UL> | ||
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Take a loopful of the slime and place it in 1 ml of sterile water in a small tube. Cap the small tube and place the capped tube into an empty 16 mm tube.<LI> | Take a loopful of the slime and place it in 1 ml of sterile water in a small tube. Cap the small tube and place the capped tube into an empty 16 mm tube.<LI> | ||
Vortex the tube to disperse the sample. (Vortexing in this way helps break up the other microbes that will be embedded in the slimy material. The other microbes are taking advantage of the by-products of Nitrogen compounds excreted by the N<sub>2</sub> fixers. ) <LI> | Vortex the tube to disperse the sample. (Vortexing in this way helps break up the other microbes that will be embedded in the slimy material. The other microbes are taking advantage of the by-products of Nitrogen compounds excreted by the N<sub>2</sub> fixers. ) <LI> | ||
Isolation streak a sample of the diluted, vortexed slime suspension following the protocol in [[BISC209: Streaking for Isolation | Streaking for Isolation ]] onto | Isolation streak a sample of the diluted, vortexed slime suspension following the protocol in [[BISC209: Streaking for Isolation | Streaking for Isolation ]] onto Azotobacteria agar medium. <LI> | ||
Incubate at room temp or at 30 °C. <LI> | Incubate at room temp or at 30 °C. <LI> | ||
'''Isolation:'''<BR> | '''Isolation:'''<BR> | ||
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Continue to isolation streak until you think you have pure isolates. <BR><BR> | Continue to isolation streak until you think you have pure isolates. <BR><BR> | ||
To test for purity from contaminants: The contaminants should be detectable on nutrient agar, so streak from an isolated colony onto nutrient agar. If more than one type of colony appears Gram stain them (because the Azotobacters | To test for purity from contaminants: The contaminants should be detectable on nutrient agar, so streak from an isolated colony onto nutrient agar. If more than one type of colony appears Gram stain them (because the Azotobacters may not appear slimy on nutrient agar) then restreak the colony with the correct morphology onto Azotobacteria medium. <LI> | ||
'''Identification:'''<BR> | '''Identification:'''<BR> | ||
Once you have pure isolates, begin to examine the cellular morphology, structure, and metabolism of your isolates as described in LAB 5.</LI></UL> | Once you have pure isolates, begin to examine the cellular morphology, structure, and metabolism of your isolates as described in LAB 5.</LI></UL> | ||
