Subtractive cDNA Cloning Using Magnetic Beads and PCR

Subtractive cloning is a method that facilitates the isolation of nucleotide sequences present in a test sample but absent or present at much lower levels, in a reference sample. A variety of situations lend themselves to this methodology and it has been used, for example, to characterize changes in levels of gene expression during development (1 –3 ), in pathological situations (4 –6 ), in inductive events (7 ,8 ), and for the isolation of tissue-specific genes (1 ,9 –11 ). Current methodologies also allow detection of pathogens and integrated retroviruses present at exceedingly low levels in the cell (12 ).