Lsm 5 pascal scanning confocal microscope

Embryos with Crk knockdown and carrying live imaging constructs were produced by crossing either crkS-RNAi/CyO; His2av mRFP, sGMCA or crkS-RNAi/CyO; MRLC::GFP males to matα tub-GAL4 II; matα tub-GAL4 III females. crkS-RNAi/ matα tub-GAL4 II; His2av mRFP, sGMCA/matα tub GAL4 III or crkS-RNAi/ matα tub-GAL4 II; MRLC::GFP/matα tub GAL4 III female progeny were then crossed to crkS-RNAi males and the progeny were imaged. CyO/matα tub -GAL4 II; His2av mRFP, sGMCA/matα tub-GAL4 III females crossed to y, w males were used as controls. Flies were allowed to lay on apple juice agar plates with yeast for 2 h. Embryos were dechorionated in 50% bleach for 3 min, washed 3× with 0.01% Triton-X, and transferred to a fresh apple juice agar plate where they were immersed in halocarbon oil 27(Apex Bioresearch Products). Seven to 10 embryos were then transferred to halocarbon oil on the gas-permeable membrane of a Lumox dish (Sarstedt) and covered with a glass coverslip. Imaging was done on a Zeiss LSM-5 Pascal scanning confocal microscope (Oberkochen, Germany). Embryos were imaged at the apical cortex where the tops of actin caps were visible. Frames were taken at 10-s intervals for ∼2 h or until cellularization began.