Question

Could someone help me with this questions. I can't really get it.

1) You are characterizing an integral membrane protein located in the plasma membrane. Based on the known amino acid sequence, you predict that the size of the protein should be 125 kDa. When you run an SDS-PAGE followed by western blotting, you find that the size is 175 kDa. What could explain the difference in predicted versus actual size of this protein?

2) If you wanted to map the chromosomal location of the HOAP gene within the Drosophila genome (i.e. determine which chromosome it resides on and where on the chromosome it is positioned), what method could you use?

Answer 1

1) The difference in size hints at some post-translational modification of the protein which couldn’t be predicted before.

2) In order to find location of gene chromosomally, one can do FISH (fluorescence in situ hybridization). Chromosomes are collected at metaphase/interphase stage. A fluorescent probe, exactly complementary to the gene of interest is then added to the chromosomal preparation. Location of fluorescence signal via microscopy would reveal the chromosome and location of gene on the chromosome.