Mapping Expressed Sequence Tags (ESTs) by Multiplexing PCR Reactions from Hybrid Cell Panels and Detecting Fluorescently Labeled Products

Determining the chromosomal origin of expressed sequence tags (ESTs) (1 ,2 ) lags far behind their identification in single-pass sequencing projects (1 –10 ). Positional cloning of disease genes requires that previously uncharacterized transcripts be mapped to the smallest possible defined region. We have developed an efficient polymerase chain reaction (PCR)-based procedure for the rapid assignment of ESTs to human chromosome regions (11 –12 ; Fig. 1 ). The critical features of the method are:

1.	Standard, restricted criteria for primer design;
2.	Sensitive, automated analysis of fluorescently labeled PCR products,
3.	Standard PCR conditions; and
4.	Multiplexed PCR reactions.

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