[35] Cross-species polymerase chain reaction: Cloning of TATA box-binding proteins

This chapter discusses an application of the polymerase chain reaction (PCR) to isolate the genomic or complementary DNA (cDNA) fragments of a gene from one species, given that the polypeptide or DNA sequence of the gene from a second species is known. The PCR technique is extremely sensitive, which means that the utmost care must be taken to avoid cross-contamination of samples, especially from micropipetting equipment. Physically separating work areas and equipment used for setting up the reactions and the subsequent analysis of the PCR products is perhaps the best way to avoid cross-contamination. The complete sequence of Drosophila and human t-butylbicyclophosphorothionates (TBPs) is analyzed in this chapter. This analysis shows that of the 10 regions of the yeast amino acid sequence chosen for designing PCR primers, three show perfect identities between the two species and other three show one amino acid sequence difference between the two species.