LAB 7: Characterization of culturable soil bacteria
Culture Independent ID of your isolates by 16S rRNA gene sequencingBR>
We have sent out all the amplicons that you prepared last week for DNA sequencing of the 16s rRNA gene. We must also send an appropriate primer. What's the sequence of such an appropriate primer? What's going on in those automatic sequencing machines in Danvers? Make sure you understand how Sanger sequencing works. We should have the data back by LAB9 when we are schedule to meet in a computer lab and learn to analyze the data generated so we can find out, we hope, a lot about the diversity of the soil bacteria in our habitat of interest.
Culturable Bacteria Characterization continued
Continue following the protocols for Tests to determine the role of a soil isolate
Concentrate on completing, reading, and assessing the tests on the isolates we sent out for anaylsis (use the appropriate protocols-refer to lab 5 and 6).
Continue with/add new differential ID test (see the appropriate protocols-refer to lab 5 and 6)
If there extra test medium is available you may repeat any tests that were ambiguous or add any you think might be useful for identifying your organism. You can also make a list of tests that would allow you to identify your isolates to species but were not available.
More adventurous students may, if you have an interesting new isolate that was not sent out for sequencing, perform tests reaction and morphology, motility, and a few simple "role" tests performed over the last few weeks. Select tests that can be completed within 2 more weeks. By this point, you should be wrapping up the testing, beginning to discard your plates and check with the instructor about freezing some of your cultures.
Be sure to fill in the data sheet found in your lab conference and enter the code for your isolate and all the morphologic observations and test results. If you are pretty sure you know the genus of an isolate, please add that as well.
1. All culture plates that you are finished with should be discarded in the big orange autoclave bag near the sink next to the instructor table. Ask your instructor whether or not to save stock cultures and plates with organisms that are provided.
2. Culture plates, stocks, etc. that you are not finished with should be labeled on a piece of your your team color tape. Place the labeled cultures in your lab section's designated area in the incubator, the walk-in cold room, or at room temp. in a labeled rack. If you have a stack of plates, wrap a piece of your team color tape around the whole stack.
3. Remove tape from all liquid cultures in glass tubes. Then place the glass tubes with caps in racks by the sink near the instructor's table. Do not discard the contents of the tubes.
4. Glass slides or disposable glass tubes can be discarded in the glass disposal box.
5. Make sure all contaminated, plastic, disposable, serologic pipets and used contaminated micropipet tips are in the small orange autoclave bag sitting in the plastic container on your bench.
6. If you used the microscope, clean the lenses of the microscope with lens paper, being very careful NOT to get oil residue on any of the objectives other than the oil immersion 100x objective. Move the lowest power objective into the locked viewing position, turn off the light source, wind the power cord, and cover the microscope with its dust cover before replacing the microscope in the cabinet.
7. If you used it, rinse your staining tray and leave it upside down on paper towels next to your sink.
8. Turn off the gas and remove the tube from the nozzle. Place your bunsen burner and tube in your large drawer.
9. Place all your equipment (loop, striker, sharpie, etc) including your microfuge rack, your micropipets and your micropipet tips in your small or large drawer.
10. Move your notebook and lab manual so that you can disinfect your bench thoroughly.
11. Take off your lab coat and store it in the blue cabinet with your microscope.
12. Wash your hands.
Study for your Lab Practical
Begin to wind down and finalize the characterization of your culturable isolates.
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