Isolated Microspore Embryogenesis in Cereals: Aspects and Prospects Archana Chugh and François Eudes Agriculture and Agri-Food Canada, Lethbridge Research Centre, 5403 1st Avenue South, PO Box-3000, Lethbridge, Alberta T1J 4B1, CANADA. Telephone: 1-403-317-3358/3338, Fax: 1-403-382-3156 e-mail:, Abstract Developmental switch of microspores towards embryogenic pathway leading to haploid plant formation, is manifestation of the dictum of ‘cellular totipotency’ at its best. Isolated microspore culture provides an excellent system to study molecular mechanism(s) of reprogramming of plant cells towards embryogenic pathway. The system also provides an alternate route to develop transgene technology in cereals that are otherwise known for their notorious recalcitrance towards various other tissue culture techniques. Exposure of microspores to a suitable stress during pre-treatment period is a pre-requisite for acquisition of embryogenic potential. As a consequence, signalling pathways associated with stress, hormone and development are activated. These pathways play key role in triggering the microspores towards embryogenesis (androgenesis). Some of the genes and proteins have been identified as molecular markers for this developmental pathway. Further improvement in the protocols for isolated microspore culture and identification of signalling molecules during various stages of haploid embryo development will open new vistas for biotechnological improvement of commercially important cereals. Isolated Microspore Culture in Cereals Anther culture is well developed in many cereals, however, it is now possible to regenerate greater number of haploid plantlets without the intervention of accessory tissues of the anther. Sheering of anthers by various methods results in isolation of microspores in the medium. Further purification step of the microspores from the anther debris makes them readily available for in vitro culture. Microspore culture generally involves the following steps: (1) Pre-treatment of the spikelet/inflorescence (with right stage of the anthers) by exposing them to a certain stress (2) isolation and purification of microspores from the pre-treated anthers and further culturing into a suitable induction medium (3) formation of multicellular structures (MCS) still contained within the exine (4) rupture of the exine and release of the MCS, which gradually develop into embryo like structures (ELS) (5) plantlet formation (6) plant maturation (Figure 1 shows steps 2-6 for isolated microspore culture in triticale, established and routinely carried in our lab). In addition, androgenesis is controlled by various environmental and physiological factors. It is highly dependent on species, genotype, the physiological condition of the donor plant, stage of the microspores, type of stress pre-treatment (temperature, starvation, osmotic stress, use of chemicals such as colchicine, recognition of suitable culture conditions for microspore induction such as ovary co-culture (Jahne and Lorz, 1995; Hu and Kasha, 1997; Reynolds et al., 1997; Touraev et al., 1997). Microspore culture is now well established for commercially important cereals such as barley (Ziauddin et al., 1992; Mordhorst and Lorz 1993; Scott and Lyne, 1994; Hoekstra et al., 1996, 1997; Kasha et al., 2001a; Li and Devaux 2001, 2003), maize (Pescitelli et al., 1990; Gaillard et al., 1991;Testillano et al., 2002), rice (Cho and Zapata 1990; Ogawa et al., 1995; Xie et al., 1995; Raina and Irfan 1998; Chowdhury and Mandal, 2001), wheat (Chu et al., 1990; Mejza et al., 1993; Hu et al., 1995; Hu and Kasha, 1997, 1999; Holme et al.,1999; Indrianto et al., 1999; Ingram et al., 1999; Liu et al., 2002a, b; Patel et al., 2004) and triticale (Keller et al., 1991; Monostori et al., 1998; Pauk et al., 2000; Oleszczuk et al., 2004; Eudes and Amundsen 2005). for further reading : Archana Chugh and François Eudes (2008) Isolated Microspore Embryogenesis in Cereals: Aspects and Prospects. In: Ashwani Kumar and Sudhir Sopory (eds) Recent advances in Plant biotechnology and its applications. IK International New Delhi pp 694