Team:KIT-Kyoto/Notebook-week6

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Revision as of 09:28, 24 September 2012






September 11th


*TNFA and API2

The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

*Parts

 pSB1C3-UAS-1, -2, -3, -4  5uL 
 3buffer(NEB)  2uL 
 BglⅡ  0.5uL 
 dH2O  12.5uL 
 Total  20uL 




 GAL4  40uL 
 4 buffer(NEB)  5uL 
 XbaⅠ(NEB)  0.5uL 
 SpeⅠ(NEB)  0.5uL 
 CIP  0.5uL 
 100×BSA  0.5uL 
 dH2O  3uL 
 Total  40uL 




 1uL 
 2.5uL 
 2.5uL 
 Ligation high  6uL 
 Total  12uL 


 2uL 
 2.5uL 
 1.5uL 
 Ligation high  6uL 
 Total  12uL 


September 12th


*TNFA and API2

The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

*Parts

 pSB1C3-UAS-1, -2  40uL 
 3buffer(NEB)  5uL 
 SpeⅠ  1uL 
 100×BSA  0.5uL 
 dH2O  3.5uL 
 Total  50uL 




 1uL 
 2uL 
 EGFP or LacZ  2uL 
 Ligation high  2.5uL 
 Total  7.5uL 


 2uL 
 2uL 
 2uL 
 Ligation high  6uL 
 Total  12uL 


September 13th


*TNFA and API2

The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

*Parts

 DNA sample  1uL 
 4 buffer(NEB)  0.5uL 
 EcoRⅠ-HF(NEB)  0.2uL 
 SpeⅠ(NEB)  0.2uL 
 100×BSA  0.05uL 
 dH2O  3.25uL 
 Total  5uL 


 DNA sample  1uL 
 3 buffer(NEB)  0.5uL 
 BglⅡ  0.2uL 
 dH2O  3.3uL 
 Total  5uL 


 1uL 
 1.5uL 
 dH2O  2.5uL 
 Ligation high  2.5uL 
 Total  7.5uL 


 2.5uL 
 2.5uL 
 2uL 
 Ligation high  7uL 
 Total  14uL 


 1uL 
 2uL 
 dH2O  2uL 
 Ligation high  5uL 
 Total  10uL 


September 14th


*TNFA and API2

The adult male and virgin female flies from microinjected embryos were mated with yw virgin female flies and yw male flies, respectively. Mated flies were transferred to the new food vials in every three days to lay eggs as much as possible. The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

*Parts

 DNA sample  1uL 
 4 buffer(NEB)  1uL 
 EcoRⅠ-HF(NEB)  0.2uL 
 SpeⅠ(NEB)  0.2uL 
 100×BSA  0.1uL 
 dH2O  7.5uL 
 Total  10uL 


 2uL 
 3uL 
 1uL(HS) or 2.5uL(Act5c) 
 Ligation high  6uL(HS) or 7.5uL(Act5c) 
 Total  12uL(HS) or 15uL(Act5c) 


September 15th


*TNFA and API2

The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

*Parts

 DNA sample  1uL 
 4 buffer(NEB)  1uL 
 EcoRⅠ-HF(NEB)  0.2uL 
 SpeⅠ(NEB)  0.2uL 
 100×BSA  0.1uL 
 dH2O  7.5uL 
 Total  10uL 




September 16th


*TNFA and API2

The adult male and virgin female flies from microinjected embryos were mated with yw virgin female flies and yw male flies, respectively. In total 83 flies from microinjected embryos were mated with yw flies. Mated flies were transferred to the new food vials in every three days to lay eggs as much as possible.

September 17th


*TNFA and API2

The progeny flies were inspected by dissecting microscope to look for the successfully transformed w+ red eye flies (red eye screening). However, no red eye fly was found.

*Parts

 DNA sample  1uL 
 4 buffer(NEB)  1uL 
 EcoRⅠ-HF(NEB)  0.2uL 
 SpeⅠ(NEB)  0.2uL 
 100×BSA  0.1uL 
 dH2O  7.5uL 
 Total  10uL