Microinjection into the lumen of the ureteric tree.
During embryonic kidney development, the ureteric bud (UB) undergoes repetitive branching to generate the entire renal collecting system. Defects in UB branching result in renal malformations, from hypoplastic kidneys to renal agenesis. Mouse genetics has become an invaluable tool to identify gene networks regulating UB branching, and the recent use of embryonic chimeras has provided further insight into the cell-autonomous regulation of this process. However, the generation of these mouse models is often resource- and time-consuming. A simplified alternative to the generation of mouse mutants or chimeras relies on the modification of UB gene expression ex vivo. This chapter describes a simple method for microinjection into the lumen of the ureteric tree of embryonic kidney explants. The mouse embryonic kidney is cultured on an air-medium interface and a thin pulled glass needle is used to access the ureteric tree and deliver the reagent of choice. The applications of the technique are multiple: from simple labeling of the ureteric tree with fluorescent markers to overexpression or downregulation of specific genes by introducing viral vectors, siRNAs, morpholinos, or other agents.
Department of Genetics and Development, Columbia University, New York, NY, USA. cc2636@columbia.edu
Cristina
C
Cebrián
D000818:Animals; D007668:Kidney / Q000196:embryology*; D051379:Mice; D008845:Microinjections / Q000379:methods*; D008853:Microscopy; D009924:Organ Culture Techniques / Q000379:methods*