In situPCR on Plant Material with Sub-cellular Resolution

In situPCR and reverse transcribedin situPCR have been testedon leaves of sugar cane fixed in FAA, 4% PFA or 2% PFA+2.5%GA.In situPCR amplification of the gene coding for ribulose-1,5-bisphosphatecarboxylase/oxygenase (rbcL) was successfully performed followingall three fixation protocols. As expected the PCR product wasrestricted to the plastids of all cells. Reverse transcribedinsituPCR was performed onrbcL mRNA and in this case the PCR productwas restricted to the plastids of the bundle sheath cells. Thisis the first report ofin situPCR on plant material and onlythe second report ofin situPCR with sub-cellular resolution.InsituPCR onrbcL may prove to be a valuable positive control forfuturein situPCR studies on plant material.Copyright 1997 Annalsof Botany Company In situPCR; reversed transcribedin situPCR; rbcL; C₄-plants