Expression profiling of glycosyltransferases using RT-PCR

The quantitative reverse-transcription polymerase chain reaction (RT-PCR) is a method used to quantify messenger RNA (mRNA) levels. It is particularly applicable to quantification of mRNAs that are transcribed at very low levels, such as glycosyltransferases (GTs). In this chapter, I describe preliminary experiments for obtaining conditions for a quantitative RT-PCR method to quantify transcript levels of GTs and related genes potentially involved in L-selectin-mediated lymphocyte homing in the mouse gastric mucosa infected by Helicobacter felis. This method was developed by modifying conventional RT-PCR protocols and does not require fluorescence-detecting thermal cyclers. The method described here is particularly useful for assaying large numbers of samples that require very accurate analysis.