Team:NYMU-Taipei/Notebook-Lab Book-fruit flies hemolymph
Lab note
Lab note
Overview
This section of our wiki contains the protocols for our insect experiments, including cultivation, infection rate tests, and hemolymph extraction.
Related experiment schedule & Lab note
Experiments
| Experiments related to B. dorsalis | Number | Date |
| Diameter of the tunnel that allows passage to one B. dorsalis | 1 | 8/30～9/1 |
| Infecting B. dorsalis with non-designed M. anisopliae | 2-1 | 9/30~10/7 |
| infecting female B. dorsalis by copulation with infected male B. dorsalis | 2-2 | 10/10~10/17 |
| Experiments related to B. dorsalis | Number | Date |
| Diameter of the tunnel that allows passage to one B. dorsalis | 1 | 8/30～9/1 |
| Infecting B. dorsalis with non-designed M. anisopliae | 2-1 | 9/30~10/7 |
| infecting female B. dorsalis by copulation with infected male B. dorsalis | 2-2 | 10/10~10/17 |
| Experiments about hemolymph extraction | Number | Date |
| Blatta lateralis | 1 | 9/20~9/30 |
| Bactro | 2 | 9/29~10/5 |
| Bombyx mori | 3 | 10/3~10/ |
| Experiments about hemolymph extraction | Number | Date |
| Blatta lateralis | 1 | 9/20~9/30 |
| Bactro | 2 | 9/29~10/5 |
| Bombyx mori | 3 | 10/3~10/ |
Routine works
| Routine tasks of feeding Bactrocera dorsalis | Date |
| Produce & change the tube with the specific agar | 8/24~9/20 |
| Produce & change water gel and the specific ratio fodder | 9/22~10/ |
| Routine tasks of feeding Bactrocera dorsalis | Date |
| Produce & change the tube with the specific agar | 8/24~9/20 |
| Produce & change water gel and the specific ratio fodder | 9/22~10/ |
| Routine tasks of hemolymph extraction | Date |
| Collect the mulberry leaves to feed the silkworm | 10/3~10/ |
| Sterilize the related equipments | 9/20~10/ |
| Routine tasks of hemolymph extraction | Date |
| Collect the mulberry leaves to feed the silkworm | 10/3~10/ |
| Sterilize the related equipments | 9/20~10/ |
Oriental fruit flies sent to lab and cultivated by large transparent box to avoid oriental fruit flies flying out.
We used both of centrifuge tube and feeding box to feed oriental fruit flies for comparing and ensuring the status of oriental fruit flies that established the independent space for observing oriental fruit flies.
B. dorsalis started to break through the cocoon. We conducted to put B. dorsalis to centrifuge tubes, avoiding the feeding box cannot load all oriental fruit flies. We poured the special agar to centrifuge tubes, and put centrifuges into entrance to catch oriental fruit flies. Then we sealed the centrifuge tubes by cotton.
(The team members put agar into centrifuge tubes which would be their feed.)
(The left picture is that oriental fruit flies broke through cocoon; the right picture is the feeding box & centrifuge tubes where fed oriental fruit flies.)
First time to Conducting Experiment 1
Changing the centrifuge tubes where oriental fruit flies lived. (Avoid the environment being mess.) /p>
Changing the centrifuge tubes where oriental fruit flies lived.
The oriental fruit flies in centrifuge tubes are dead quickly, so we conducted trouble shooting and sent the mail to professors who research oriental fruit flies.
We took the trap to Beitou orchard to try to catch oriental fruit flies.
Searching the store selling the silkworm for extracting hemolymph which is used to induce pMcl1.
Visiting two farmers working in the orchard to ask the information about oriental fruit flies in the nature environment, and installing the trap of catching oriental fruit flies.
(Left picture: the team member install the trap; Right picture: the farmer and our team member took a picture)
Sent the mail to researchers to ask for making public the content of the mail.
Communicating with Chang Ming ecological education sericulture, asking the information of buying silkworm.
Picking up the oriental fruit fly larval from rotten guavas and feeding them.
Left picture: the team members were picking up the oriental fruit fly larval from rotten guavas; Right picture: the larval have picked up and put into another box. Expected becoming pupae after six days.
For the situation that our fruit flies in the centrifuge tubes were dead quickly , so we visited Dr. Yu-Bing Huang in Taiwan Agricultural Research Institute, Applied Zoology Division, and asking the problems about the right way for feeding oriental fruit flies. We change our way of feeding oriental fruit flies.
Producing new feeding box which is larger and ventilation, and design the experimental boxes for Experiment 2: Infecting B. dorsalis with non-designed M. anisopliae.
(Left picture: experimental boxes; Right picture: the internal space of feeding box)
The new oriental fruit flies broke through cocoon in the new feeding box
the new way for feeding oriental fruit flies is using gauze to be the windows that could be ventilation, and we can put watered agar and artificial feed on the top of gauze, then there were no water and feed in the box which lead to the environment being mess, and they also could have enough water and feed.
Infecting B. dorsalis with non-designed M. anisopliae started to conduct.
Photos shows three concentration condition and negative control environment of experiment "Infecting B. dorsalis with non-designed M. anisopliae"
10/2: GFP_TtrpC digested for ligation with pBARGPE1
10/2: Transformed pBAR_GFP_TtrpC into competent cells
10/2: pBARGPE1 digested for pBAR_GFP_TtrpC construction
10/2: pBARGPE1 resuspended (from cryopreservation)
10/3: Extracted plasmid pBARGPE1
10/3: pBAR_GFP_TtrpC containing cultures used in 2 in 1
10/3: Extracted plasmid pBAR_GFP_TtrpC
10/3: pBAR_GFP_TtrpC digestion check(failure)
10/3: pBAR_GFP_TtrpC PCR check(failure)
10/3: Transformed pBAR_GFP_TtrpC into competent cells
10/3: pBARGPE1 checked with restriction digestion (correct)
10/4: pBARGPE1 plasmid stored
10/4: pBAR_GFP_TtrpC containing cultures used in 2 in 1
10/4: pBAR_GFP_TtrpC plasmid extraction
10/4: pBAR_GFP_TtrpC digestion check
