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Lab Meeting
Bo Shi2015-09-22
Outlines
• 1 Dpr and DIP primer testing
• 2 Confocus imaging
• 3 Single cell dissociation
• 4 Transgenetic flies (just starting)
• 5 Dprs and DIPs recombining flies (just starting)
1 Dpr and DIP primer testing
• The primers of 12 Dprs have been tested, and can be used for the next step. (The other 8 or 20 Dprs has been tested before my last lab meeting.)
• 10 of 11 DIPs also can be used. (exception: CG40378)
2nd round PCR imagesNo. 1 2 3 4 5 6
Genes Dpr1 Dpr 3 Dpr 7 Dpr 8 Dpr 9 Dpr 10-new
cDNA 201 248 270 220 457 225
gDNA N/A N/A N/A 8661 5056 888
1 2 3 4 5 6
No. 1 2 3 4 5 6
Genes Dpr11-new
Dpr15-new
Dpr16-new
Dpr17-new
Dpr 19 Dpr 20
cDNA 242 208 370 273 X X
gDNA 826 3500 801 2124 X X
1 2 3 4 5 6
No. 1 2 3 4 5 6
Genes Dpr19-new
Dpr 20-new
DIP-α(1) DIP-γ(3) DIP-δ(4) DIP-ε(5)
cDNA 202 337 288 385 318 311
gDNA N/A N/A 1332 841 1732 5272
1 2 3 4 5 6
No. 1 2 3 4 5 6
Genes DIP-ζ(6) DIP-η(7) DIP-θ(8) DIP-ι(9) CG31814 CG40378
cDNA 274 X 320 274 253 X
gDNA N/A X N/A N/A 3225 X
1 2 3 4 5 6
No. 1 2 3
Genes DIP-β(2) DIP-η(7)-new
CG40378-new
cDNA 309 /224 324 X
gDNA 15049 N/A X
1 2 3
2 Confocus imagingort-c1a-lv
Tm20: YW ; 17D01-Flp
;lot-stop-mCD8-GFP
+
Tsc mutation, Tm20 ;♂ 122 ; C1-Gal4, UAS-mCD8-Cherry
+
FRT82B, Gal80
FRT82B, Tsc1
3 Single cell dissociation
♀ YW ; 17D01-Flp
Cyo;
+
Tm2 or Tm6BX ♂ YW ;
ort-c1a-lv
Cyo;
lot-stop-mCD8-GFP
Tm2 or Tm6B
ort-c1a-lvF1: YW ;
17D01-Flp;
lot-stop-mCD8-GFP
+
FH1
2nd round PCR imagesmCD8 1 2 3 4 5 6 7 8
vGlut 1 2 3 4 5 6 7 8
Primers cDNA gDNA
mCD8 266 266
vGlut 339 807
4 Transgenetic flies
• Rh3-syb-spGFP1-10• Rh4-syb-spGFP1-10• Rh5-syb-spGFP1-10• Rh6-syb-spGFP1-10• UAS-YFP-sp1-10cyto
5 Dprs and DIPs recombining flies
Dpr1 2R 57B1-57B2 FRT42
Dpr20 3L 61D2-61D2 FRT2A
Dpr3 2L 22C1-22C3 FRT40
CG14010 2L 25F1-25F1 FRT40
CG31708 2L 29F8-30A1 FRT40
Future Work
• 1 Complete CG40378 primers design and test.• 2 Test DIPs and Dprs expression in the medulla.• 3 Test DIPs and Dprs expression in Tm20, Dm8 &
Tm5C.• 4 Inditify which Ach receptor expresses in
Tm20.• 5 Screen some adhesion molecule mutations in
Tm20.
• 6 Continue to pick Transgenetic flies• 7 Continue to recombine Dprs and DIPs flies
with FRT flies• 8 Obtain the specificity of Tm20 in the line of
ort-c1a-lvF1: YW ;
17D01-Flp;
lot-stop-mCD8-GFP
+
Transgenetic flies X YW
F1: 1 red eye fly♂ X YW; ;Cyo Tm6B
Tm2B13♀
If transgne is on the ChX, all females are red eyes and males are white eyes in the F2 geneation.
If Transgene is on the on the Ch2, Ch3 or Ch4, then:
F2: 1 red eye fly♂ X YW ; ;Cyo Tm6B
Tm2B13♀
3 Single cell dissociation
♀ YW; 17Dol-Flp
Cyo;
+
Tm2 or Tm6BX ♂ YW;
ort-cla-lv
Cyo;
lot-stop-mCD8-GFP
Tm2 or Tm6B
;♂ 122 ; C1-Gal4, UAS-mCD8-Cherry
+
FRT82B, Gal80
FRT82B, Tsc1
