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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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When the solution begins to look turbid, slowly loosen the cap while watching for the appearance of bubbles rising in the liquid. The bubbles appear because of decreased pressure from loosening the cap and indicate the production of N<sub>2</sub> gas by the organism. <LI> | When the solution begins to look turbid, slowly loosen the cap while watching for the appearance of bubbles rising in the liquid. The bubbles appear because of decreased pressure from loosening the cap and indicate the production of N<sub>2</sub> gas by the organism. <LI> | ||
Make a wet mount of the culture by taking a loop of liquid (after scraping the sides of tube with your inoculating loop). This scraping will help you collect prosthecate as well as motile cells.<LI> <UL><LI> | Make a wet mount of the culture by taking a loop of liquid (after scraping the sides of tube with your inoculating loop). This scraping will help you collect prosthecate as well as motile cells.<LI> <UL><LI> | ||
http://www.youtube.com/watch?v=ZMiWdOl_9i0&feature=related YouTube link to making a wet mount slide]<UL><LI> | |||
Take precautions when handling the slides.<LI> | Take precautions when handling the cultures and slides.</LI> | ||
<ul><LI> | |||
If your organism is growing in a broth, aseptically use your loop to place a loopful of broth directly on the slide. If you are making a wet mount from a solid colony on a plate, place a small drop of water on a slide and use your loop to aseptically transfer a barely visible amount of bacterial growth into the water drop and gently swirl. Do not spread the drop.<LI> | |||
Place a coverslip at a 45 degree angle on the edge of the drop of water and release it gently to spread the drop under the coverslip.<LI> | Place a coverslip at a 45 degree angle on the edge of the drop of water and release it gently to spread the drop under the coverslip.<LI> | ||
With the diaphragm closed to reduce the light as much as possible, | With the diaphragm closed to reduce the light as much as possible, use the 10x objective to locate the edge of the coverslip (a big black vertical line). Get that in sharp focus and then move the viewing field just slighty into the water drop. Locate the interface between the air and edge of teh water drop (a thin black line). Get that in sharp focus and then move to the 40x objective and using only the fine focus on the water side of the drop you should see the bacteria. DO NOT USE THE OIL IMMERSION LENS FOR A WET MOUNT. The organisms will be very small, due to the low magnification. Often the highest concentration of organism are seen along the edge of the drop.<LI> | ||
Do not discard this slide, full of living organisms, in the autoclave bags or the glass disposal box. Discard the slide in the container of disinfectant located beside the sink near the instructor desk. </LI></UL></LI></UL> | |||
This technique is difficult, please contact your instructor for help if you are unable to find the edge of the water drop. Once you are successful, you will find this technique extremely useful to quickly check the morphology of your bacterial isolates. | |||
Discard the slide in the disinfectant container, not the glass disposal box.</LI></UL> | Discard the slide in the disinfectant container, not the glass disposal box.</LI></UL> | ||
