Lab 7

Using Fluorescence microscope to observe the Neurospora’s protein localization & observe Chlorophyllum sp. mushroom’s clam connection and Buller’s drop

Objectives: 1) Learn how to use fluorescence microscope to locate the proteins which tagged with fluorescence proteins, and familiar with procedure of fluorescence microscope  2) Observe the clam connection and Buller’s drop from Chlorophyllum sp. which are two features of mushroom.

Materials and Methods

1: N. crassa SMRP10 x CSP1-GFP 

2: Field-collected Chlorophyllum sp. mushroom

3: Water agar medium in Petri dishes

4: A kit containing microscope slides, cover slips, needles, and transfer loops.

5: Immersion oil and dropper bottles for water to suspend specimens. 

6: Lens paper for cleaning objectives and Kim wipes for working with microscopic slides and specimens.

 7: Bunsen burner, dissecting needle.

8: Microscopy: Canon Powers hot SD550 digital camera
Olympus SZ30 zoom stereo microscope  Olympus CX31 compound microscope

Procedures:

1: Make the mount slides. Cut the fungi culture with the agar from the petri dish, and place on slide. Observe the fluorescence by microscopy (The produces was performed by Dr. Shaw).

3: Observe the protein localization by fluorescence microscopy.

Observations:

1)

Fig 1. 2. 3. Danimic of actin-GFP protien in N.crassa

II)

                                        Fig. 1. Chlorophyllum sp. mushroom from field

                                   

                                      Fig. 2, 3. Spores were released from gill

                                       Fig. 4. Clam connetion

Discussion:

Failed to find very obviously growing hyphae, however, we observed the fluorescence proteins were under the dynamic movment in the  hyphae.

The clam connetion was obsereved under the light microscopy, it is like a bridge to link two cells.  We failed to find the Buller's drop from this sample, may be the way we prepared the sample is too harsh that have destoryed this stucutre, or it is possible that the mushroom is at the not the right status which produce this stucutre.