Imaging Migrating Border Cells in Drosophila Ovaries

Male flies containing slbo-GAL4, UAS-lifeact-GFP and UAS-LacZ were crossed to UAS-raskol-RNAi females. 1–2 day old F1 females were incubated at 29°C for 1–2 days in vials with fresh yeast paste and slbo-GAL4, UAS-lifeact-GFP males. Ovaries were dissected for live imaging in imaging media (Schneiders’s medium, 15% fetal bovine serum (FBS) and 0.2mg/ml Insulin; Thermo Fisher Scientific) according to published protocols [22 (link), 98 (link)]. 100 μl of imaging media containing egg chambers was transferred to poly-D-lysine coated Mattek dishes for imaging. 20 μm z-stacks (1 μm step size) covering the whole migrating border cell cluster were acquired every 2 minutes using on a Nikon A1 scanning confocal microscope.

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Raza Q., Choi J.Y., Li Y., O’Dowd R.M., Watkins S.C., Chikina M., Hong Y., Clark N.L, & Kwiatkowski A.V. (2019). Evolutionary rate covariation analysis of E-cadherin identifies Raskol as a regulator of cell adhesion and actin dynamics in Drosophila. PLoS Genetics, 15(2), e1007720.

Publication 2019

Insulin A1 scanning confocal microscope

Cell Confocal microscope Dishes Females Fetal bovine serum Flies Insulin Lacz Lysine Males Ovaries Paste Poly Rnai Yeast

Corresponding Organization : New York University

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Variable analysis

independent variables

Presence of UAS-raskol-RNAi

dependent variables

Behavior of the migrating border cell cluster

control variables

Presence of slbo-GAL4
Presence of UAS-lifeact-GFP
Presence of UAS-LacZ
Incubation at 29°C for 1-2 days
Use of fresh yeast paste
Presence of slbo-GAL4, UAS-lifeact-GFP males
Use of imaging media (Schneiders's medium, 15% fetal bovine serum (FBS) and 0.2mg/ml Insulin)
Poly-D-lysine coated Mattek dishes for imaging
Acquisition of 20 μm z-stacks (1 μm step size) every 2 minutes using a Nikon A1 scanning confocal microscope

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