Whole-mount in situ hybridization detection of mRNA in GFP-marked Drosophila imaginal disc mosaic clones.
Protocols for both generation of mosaic clones and wholemount in situ hybridization detection of mRNA have led to an advanced understanding of Drosophila development. A simple means of combining these techniques for staining of imaginal discs would be useful, as it would allow for analysis of mRNA transcripts in marked mosaic clones. However, few researchers attempt such experiments due to the technical difficulty of simultaneously detecting clones and mRNA transcripts. Furthermore, maintaining the ability to use GFP-marked clones is desirable. However, typical Drosophila in situ hybridization protocols result in loss of GFP fluorescence. The method for double labeling of imaginal discs described here maintains the ability to identify mosaic clones, including those marked by GFP, while simultaneously allowing the detection of mRNA transcripts. The methodology described is technically manageable, robust, rapid, relatively inexpensive and of interest to many Drosophila researchers.
VanZomeren-Dohm
Adrienne
A
Indiana University School of Medicine-South Bend at Notre Dame, South Bend, Indiana, USA.
Flannery
Ellen
E
Duman-Scheel
Molly
M
D000818:Animals; D002999:Clone Cells / Q000378:metabolism; D004330:Drosophila / Q000196:embryology*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D017403:In Situ Hybridization / Q000379:methods*; D009030:Mosaicism / Q000196:embryology; D012333:RNA, Messenger / Q000032:analysis*
D012333:RNA, Messenger; D049452:Green Fluorescent Proteins