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Additional index words. adventitious roots, anatomy, Carya illinoinensis, embryo culture, in vitro Abstract. The regenerative capacity of mature pecan (Carya illinoinensis (Wangenh.) K. Koch) embryonic tissues was demonstrated after pretreating mature nuts to eliminate associated endogenous contaminants. Cultured cotyledon segments were induced to form adventitious roots in a medium with 50 µM NAA. A regeneration medium with 20 µM BA and 5 µM IBA stimulated prolific axillary shoot production from the embryonic axis without causing cotyledon abscission. Cotyledon retention was essential for shoot initia- tion and long-term development. Eighty-five percent of the shoots emerging from embry- onic axes formed at the cotyledonary nodes. Thirty percent of the microshoots rooted on an auxin-free medium after preculture in a medium with 20 µ M IBA. TDZ (25 µM) stimulated callus production from the cotyledonary nodes and radicles. Adventitious buds emerged on the callus surface and internally in callus. Chemical names used: a -naphthaleneacetic acid (NAA); 6-benzylaminopurine (BA); indole-3-butyric acid (IBA); N-phenyl-N'-1,2,3-thidiazol-5-ylurea (TDZ). Developing efficient, practical pecan re- eliminated by sterilization procedures without generation strategies would broaden the op- disturbing explant viability (Obeidy and Smith, portunities for in vitro germplasm manipula- 1990). In this study, we used a medium with tion and expedite advances in crop improve- low water availability to yield contaminant-free ment research. However, several authors have embryo explants from mature nuts. From these reported that only immature pecan explants explants, we successfully regenerated roots respond to in vitro culture. Proliferation using from mature pecan cotyledons and proliferated pecan microcuttings derived from seedlings shoots and roots from mature zygotic embry- has been attempted, but shoot formation has onic axes. been infrequent (Hansen and Lazarte, 1984; Mature, dormant 'Stuart' pecan nuts were Wood, 1982). Somatic embryogenesis has been purchased from a commercial sheller (Peebles accomplished only from immature zygotic Pecan Co., Starkville, Miss.) and stored for at embryos of several cultivars (Merkle et al., least 30 days at 4C before being used. Unshelled 1987; Wetzstein et al., 1988, 1989; Yates and nuts were treated with 95% ethanol for 5 min, Reilly, 1990). Various research groups have then with a 5.25% sodium hypochlorite solu-

Organogenesis from Mature Pecan Cotyledons and Embryonic Axes