Posted: March 8th, 2016

Analysing Gene function – practical molecular biology

8) Consider Fig. 4, expression of WT1 mRNA in the developing human kidney. Panels A and B show low power views of sections through the whole kidney, whilst C-H show high power views of three stages in glomerular development. By reference to the text of the original paper (Mundlos et al, 1993, Development 119, 1329-1341), state in which cell types WT1 mRNA is expressed. (2 marks)
Antibody staining
Antibodies specific to particular proteins can be used to stain whole embryos or sections of tissue to examine where the protein is expressed. Fixed embryos or sectioned material is incubated with the antibody under conditions where the specific antigen are recognised, excess unbound antibody is washed off, and bound antibodies are detected using enzymatic labels.

9) Consider Fig. 5, WT1 protein expression in the developing human kidney. Is the expression of WT1 protein in the three stages of glomerular development consistent with the expression pattern of WT1 mRNA shown in Fig. 4? (1 mark)
10) Consider Fig. 6, WT1 protein expression in the adult and fetal podocytes. The figure shows that WT1 protein is localised to the nucleus of these cells. What is one advantage of antibody staining over in situ hybridisation? Is this consistent with what you know of the WT1 gene sequence? (3 marks)