Difference between revisions of "Team:NYMU-Taipei/Notebook-Lab Book-fruit flies hemolymph"
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| − | <li><p style="font-size: 18px;"> | + | <li><p style="font-size: 18px;">Picking up the oriental fruit fly larval from rotten guavas and feeding them.</p></li> |
| − | <li><p style="font-size: 18px;"> | + | <li><p style="font-size: 18px;">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. </p></li> |
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Revision as of 19:33, 17 October 2016
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.
9/11: Standard part PgpdA amplification PCR with KOD polymerase
9/11: pBARGPE1 digested for pBAR_PgpdA_EL222_TtrpC construction
9/11: TtrpC KOD PCR amplification
9/11: TtrpC KOD PCR amplification
9/11: pBAR_PgpdA_EL222_TtrpC containing culture used in 3 in 1
9/11: pBAR_KR transformed into competent cell
9/11: pBAR_KR transformed into competent cell
9/12: pBAR_KR performed plasmid extraction and plasmid PCR (successful)
9/12: pBAR_PgpdA_EL222_TtrpC checked with restriction digestion
9/12: pBAR_PgpdA_EL222_TtrpC amplified with plasmid PCR
9/12: Successfully constructed the plasmid pBAR_PgpdA_EL222_TtrpC
9/12: Standard part PgpdA mutation PCR (mutated PstI)
9/12: Standard part PMCL1 amplification PCR with KOD polymerase
9/13: Standard Part PMCL1 mutation PCR (mutated SpeI)
9/13: Standard part TtrpC amplification PCR with KOD polymerase
9/13: pBAR _RFP_TtrpC transformed into competent cell
9/13: RFP_TtrpC PCR to amplify the target sequence
9/13: pBAR _RFP_TtrpC did 3 in 1
9/13: pBAR _RFP_TtrpC digestion check
9/14: pBAR _RFP_TtrpC plasmid extraction
9/14: RFP_TtrpC digested
9/14: K118400 and TtrpC used PCR to amplify the target sequence
9/14: KR_TtrpC used PCR to amplify the target sequence
9/15: KR_TtrpC digested with restriction enzymes
9/15: Standard part TtrpC mutation PCR (mutated XbaI)
9/15: pBARGPE1 digested for pBAR_KR_TtrpC construction
9/16: pBAR_KR_TtrpC transformed into competent cell
9/16: Transformed pBAR_KR_TtrpC into competent cell
9/16: Digested pBAR_KR for transforming into Metarhizium anisopliae
9/16: Extracted plasmid BBa_K118400
9/16: pBAR_EL222_TtrpC did plasmid PCR
9/16: pBAR_EL222_TtrpC did plasmid PCR (confirmed successfully)
9/16: pBAR _RFP_TtrpC did digestion check
9/17: pBAR_RFP_TtrpC did digestion check
9/17: pBAR_EL222_TtrpC plasmid stored and transformed cultures cryopreserved
9/17: pBAR_KR_TtrpC did 3 in 1
9/17: Extracted plasmid pBAR_KR_TtrpC
9/18: Transformed pBAR_RFP_TtrpC into competent cell
9/18: pBAR_RFP_TtrpC did 3 in 1
9/18: pBAR_KR_TtrpC did plasmid PCR and digestion check
9/18: BBa_E0040 did plasmid PCR
9/19: Amplified GFP from plasmid BBa_E0040
9/19: Extracted plasmid BBa_E0040
9/20: KR_TtrpC digestion
9/20: Transformed pBAR_KR_TtrpC into competent cell
9/20: pBAR_KR_TtrpC did 3 in 1
9/20: Extracted plasmid pBAR_KR_TtrpC
9/20: pBAR_KR_TtrpC did digestion check
9/20: Extracted plasmid pBAR_RFP_TtrpC
9/20: pBAR_RFP_TtrpC did digestion check
9/20: RFP_TtrpC digestion
9/21: pBAR_KR_TtrpC did digestion check
9/21: pBAR_RFP_TtrpC did digestion check
9/21: pBAR_RFP_TtrpC did 2 in 1
9/21: pBARGPE1 and RFP_TtrpC digestion
9/22: Amplified RFP_TtrpC with KOD polymerase
9/22: Extracted plasmid pBAR_KR_TtrpC
9/22: pBAR_KR_TtrpC did plasmid PCR
9/22: pMCL1 and pBAR_KR_TtrpC digestion (pMCL1 was not digested successfully)
9/22: pBAR_PgpdA_EL222_TtrpC digested for transforming into Metarhizium anisopliae
9/23: Amplified pMCL1 with KOD polymerase
9/24: pBAR_RFP_TtrpC did digestion check
9/24: Extracted plasmid pBAR_RFP_TtrpC (from 9/5 plate)
9/25: Transformed pBAR_RFP_TtrpC into competent cell
9/25: RFP_TtrpC digestion
9/25: Amplified RFP-TtrpC with KOD polymerase
9/25: Amplified RFP_TtrpC with KOD polymerase
9/25: pMCL1 and pBAR_KR_TtrpC digestion
9/25: Transformed pBAR_pMCL1_KR_TtrpC into competent cell
9/25: Transformed pBAR_pMCL1_KR_TtrpC into competent cell
9/25: pBAR_pMCL1_KR_TtrpC did 3 in 1
9/26: Extracted plasmid pBAR_pMCL1_KR_TtrpC
9/26: pBAR_RFP_TtrpC did 2 in 1
9/26: Extracted plasmid pBAR_RFP_TtrpC
9/26: pBAR_RFP_TtrpC did digestion check
9/26: pBAR_RFP_TtrpC did digestion check
9/26: Amplified pMCL1 with KOD polymerase
9/27: Amplified pMCL1 with KOD polymerase
9/28: pBAR_pMCL1_KR_TtrpC did digestion check
9/28: pBAR_RFP_TtrpC did plasmid PCR
9/28: Successfully constructed the plasmid pBAR_RFP_TtrpC
9/29: Amplified pMCL1 with KOD polymerase
9/30: pBAR_pMCL1_KR_TtrpC did digestion check
9/30: Received the junction primer of GFP-TtrpC
9/30: GFP_TtrpC ran ligation PCR
10/1: GFP_TtrpC ran ligation PCR
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
