Comparison of mRNA expression between wildtype and Wnt9bcneo/cneo E15.5 urogenital systems.

Wnt9b is expressed in the ureteric bud of the kidney at all stages of development. The Wnt9b cneo allele functions as a partial loss of function. Wnt9bcneo/cneo mutant kidneys initially develop normally but exhaust their nephron progenitor cells by E15.5. Here, we have compared expression between Wnt9bcneo/+ and Wnt9bcneo/cneo kidneys. Additional urogenital tissue (adrenal glands, reproductive tracts and bladder) may have been included in some samples. We used microarrays to compare differences in gene expression between wildtype and Wnt9b mutants. To focus on direct targets, mesenchymes isolated away from the E11.5 ureteric bud were assayed Ell.5 kidneys were manually dissected away from embryos collected from Wnt9b+/- intercrosses. Mesenchymes were isolated away from the ureteric bud after a brief enzymatic digestion (trypsin/pancreatin). Paired mesenchymes were stored int RNA-Later until the genotype was determined. 25-30 Wnt9b-/- or Wnt9b+/- mesenchymes were pooled for each replicate (giving approx. 1ug of total RNA). cRNA was synthesized and hybridized to the Affymetrix GeneChip Mouse Genome 430.2 Array.