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====Imaging==== *3 Tries *#Add FISSEQ_Adpt_Cy3 and then add focusclear for 2 hrs *#Add FISSEQ_Adpt_Cy3 and then 5uM DRAQ5 and then add focusclear for 2 hrs *#Embed in 5% 199:1 polyacrylamide gel and then add FISSEQ_Adpt_Cy3 #Try 1 #*Preheat 0.5uM FISSEQ_Adpt in 30% formamide + 2X SSC to 75C #*Add 100ul to embryo in tube and let sit for 10min at RT #*Wash with 2X SSC twice #*Add Focusclear and incubate at RT for 2hr #*Mount on slide with Mountclear #Try 2 #*Preheat 0.5uM FISSEQ_Adpt in 30% formamide + 2X SSC to 75C #*Add 100ul to embryo in tube and let sit for 10min at RT #*Wash with 2X SSC twice #*Add 100ul 5uM DRAQ5 in 1X PBS and let sit for 15min at RT #*Remove DRAQ5 #*Add Focusclear and incubate at RT for 2hr #*Mount on slide with Mountclear #Try 3 #*Embed in 5% 199:1 Acr:Bis Hydrogel on #1.5 Vectabond treated coverslip #**Follow standard protocol, polymerize in 50uM spacer jig for 30min in Argon filled bag #*Attach coverslip to bottom of dish #*Wash with 1X PBS twice #*Preheat 0.5uM FISSEQ_Adpt in 30% formamide + 2X SSC to 75C #*Add 100ul to embryo in tube and let sit for 10min at RT #*Wash with 2X SSC twice #*IMAGE #*Add Focusclear and incubate at RT for 2hr #*IMAGE
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