plant regeneration

callus induction

leaf explants

vitro propagation

indole butyric acid

shoot proliferation

micropropagation

plantlets

liquid medium

naphthaleneacetic acid naa

In Vitro Cellular & Developmental Biology - Plant

Black walnut ( Juglans nigra L.) has long been prized for its timber, leading to commercial cultivation and significant breeding efforts for improving marketable traits. Vegetative and in vitro black walnut propagation techniques, however, are variable and highly genotype dependent. Optimizing plant growth regulator type and concentration are integral for developing a successful micropropagation protocol. The addition of meta-topolin (MT) combined with the novel use of a liquid medium has led to the development of a rapid shoot multiplication system. The objective of this research was to develop a reproducible and dependable micropropagation protocol for elite black walnut genotypes. In vitro shoot cultures were established from nodal explants cultured on semi-solid Driver and Kuniyuki walnut (DKW) medium supplemented with 8.9 μM benzyladenine, 0.005 μM indole-3-butyric acid (IBA), 200 mg L −1 casein hydrolysate, 50 mg L −1 adenine hemisulfate, 2 mL L −1 Plant Preservative Mixture™, and 4.1 μM MT. Long-term survival and proliferation of microshoots was achieved when nodal segments of in vitro grown shoots were cultured in liquid initiation medium in 3-L polycarbonate Fernbach-style flasks on a rotary shaker (100 rpm) under a 16-h photoperiod at 25°C. Elongated microshoots (5–7 cm in length) were rooted in a slurry-like medium composed of half-strength DKW medium with 0.11% ( w/v ) Phytagel™ and coarse vermiculite (2:1, v/v ) supplemented with 50 μM IBA for 5 wk. Rooted shoots were acclimatized to ambient culture room conditions, but plantlets did not survive once transferred to the greenhouse.

Rapid in vitro shoot multiplication of the recalcitrant species Juglans nigra L.