J. Plant Physiol. 158. 1089–1092 (2001)  Urban & Fischer Verlag http://www.urbanfischer.de/journals/jpp Short Communication Single cell analysis technique for comparison of specific mRNA abundance in plant cells Joe A. Gallagher 1 * , Olga A. Koroleva 2 , Deri A. Tomos 2 , John F. Farrar 2 , Christopher J. Pollock 1 1 Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, Ceredigion SY23 3EB, UK 2 School of Biol. Sciences, University of Wales, Bangor, Gwynedd, LL57 2UW, UK Received January 29, 2001 · Accepted January 29, 2001 Summary A single cell sampling technique for determining changes in gene expression in a range of cell types is described. Expression of actin and changes in expression of a light inducible gene (sucrose : fructan fructosyltransferase) was demonstrated in mesophyll and parenchymatous bundle sheath cells. This method provides a reliable technique for the analysis of specific gene transcript abun- dance in individual plant cells. Key words: Hordeum vulgare – Lolium temulentum – micro-sampling – single cell analysis Abbreviations: RT-PCR reverse transcription-polymerase chain reaction Introduction Plant leaves are highly heterogeneous with respect to metab- olism, enzymology, and gene expression. Differences in anat- omy, function, enzyme activity, and solute composition have been demonstrated for several cell types. Variable distribu- tion of starch, sucrose, hexoses, and fructan between meso- phyll and parenchymatous bundle sheath has been found in cereal leaves (Williams et al. 1989, Koroleva et al. 1998). Het- erotrophic epidermal cells have been shown to have different patterns of both metabolism and enzyme activity when com- pared to other leaf cells (Williams et al. 1989, Obenland et al. 1993, Tomos et al. 1992, Fricke et al. 1994, Koroleva et al. * E-mail corresponding author: joe.gallagher@bbsrc.ac.uk 1997, 1998). Vascular tissue has also been shown to be differ- entiated in terms of distribution of key enzymes (Kingston- Smith and Pollock 1996). In addition, metabolic hetrogenicity has been observed among the photosynthetic cells, which constitute only about 55 % of the volume of the mature cereal leaf (Jellings and Leech 1982). These observations emphasise the importance of studying gene regulation at the cellular level. A number of techniques have been reported for measuring gene expression at the level of the single cell. These include in situ hybridisation (Langdale 1994), in situ RT-PCR (Matsuda et al. 1997), RT- PCR following micro-sampling (Brandt et al. 1999), con- struction and amplification of cDNA libraries following either single cell aspiration (Karrer et al. 1995), or isolation of tissue specific protoplasts (Dresselhaus et al. 1994, Richert et al. 0176-1617/01/158/08-1089 $ 15.00/0