PROPAGATION AND RETESTING OF WALNUT ROOTSTOCK GENOTYPES PUTATIVELY RESISTANT TO PESTS AND DISEASES Wesley P. Hackett, Gale McGranahan, Bruce D. Lampinen, Chuck Leslie, Greg Browne, Diego Bujazha, and Soussan Hirbod ABSTRACT Establishment of the field stock block of putatively disease and pest resistant rootstock genotypes was completed with the exception of 5 WIP genotypes tolerant to CLRV that will be grafted in 2004 to already established California black walnut rootstocks. The tissue culture bank of high priority genotypes has been completed with the exception of 87-32-1 (WIP5), a CLRV tolerant genotype. We have continued work to micropropagate the disease and pest resistant genotypes using improvements to the fog chamber acclimatization and greenhouse growing protocols developed during Year 1 of the project. Through October 2003, we have micropropagated over 5000 plantlets of 20 genotypes in 1½ inch tree tubes. We have shown that we can keep these plantlets growing year round in a greenhouse using a daylength extension with HID lights and a Promalin spray treatment every 2 weeks. We have also shown that plantlets in 1 1/2 inch tree tubes can be greenhouse grown to a size (4-5 inches tall with 3-5 tap roots 6 inches long) large enough for transplanting to a nursery row in 3 months. Three-month-old plants can be can be induced to drop their leaves by placing them in cold storage with low light at 50 F for 3 weeks. These dormant plantlets can then be stored for up to 6 months in a cold room at 42 F. Alternatively, they can be placed outdoors in fall and winter for dormancy induction and breaking. Rooting of microshoots is the main factor that limits successful micropropagation. Some genotypes root very well (60-70%) while other root considerably below the average (40%). Some genotypes recalcitrant for rooting responded well to higher IBA concentrations than had previously been used while others did not. However, use of higher IBA concentrations appears to be a promising way to obtain better rooting of many of the genotypes. For the period November 2002 to October 2003, the average survival of all genotypes was 65% of lab-rooted plantlets that were transplanted to 1½ inch tree tubes with considerable variation between genotypes. A total of 1300 hardwood cuttings were taken from 10 coppiced or tree form genotypes. Rooting percentage varied from 0-100%. Genotype 87-32-1 (WIP5, one which we have trouble micropropagating), a CLRV tolerant clone, rooted at nearly 100% and VX 211, a nematode tolerant genotype rooted at 87%. Rooting of hardwood cuttings also showed some potential (35-50% rooting) for 87-50-1 (WIP6), 87-27-4 (WIP3), 87-27-6 (WIP4) and AZ025, a crown gall resistant genotype for which we have trouble micropropagating. Using methods referred to above, we now have a total of over 3900 trees of 21 genotypes available for field trials in 2004. We have supplied 493 plantlets of 7 genotypes in 1½” inch tree tubes to Greg Browne for Phytophthora retests during Year 2 and also have 407 plantlets of 8 genotypes in cold storage for addition Phytophthora retests during 2004. Genotype RX1 was relatively resistant to P. citricola in two 2003 experiments, confirming previous results with the clone. Imposing a cycle of pre-inoculation chilling, dormancy, and growth resumption on the genotypes before screening them facilitated assessment of their genetic susceptibility to the pathogen.