A two-step pretreatment significantly enhances shoot organogenesis from leaf explants of highbush blueberry cv.Bluecrop

As part of a program to improve highbush blueberry (Vaccinium corymbosum L.) cultivars via tissue culture and genetic engineering, studies were conducted to determine optimum conditions for organogenesis from leaf explants of the previously recalcitrant cv. Bluecrop. The effects of a pretreatment, growth regulators, and age of explant source on shoot organogenesis were investigated. A maximum of 98% explants regenerated shoots with a mean of 11 shoots per leaf explant after 62 days when explants of 2-week-old shoot cultures were incubated on the following regime: pretreatment medium #1 containing 5 muM TDZ and 2.6 muM NAA for 4 days, pretreatment medium #2 containing 7 muM zeatin riboside and 2.6 muM NAA for 3 days, regeneration medium containing 1 muM TDZ for 6 weeks, and last on medium without growth regulators for 10 days. No shoot regeneration occurred if explants were incubated without exposure to pretreatment prior to incubation on regeneration medium. There were no significant differences in percentage of regeneration or the number of shoots regenerating per explant from leaf explants derived from either 1-, 2-, or 3-week-old shoot cultures. Shoot production per explant on regeneration medium containing 1 muM TDZ was about three times that on 0.5 muM TDZ or 20 pm zeatin riboside, and nine times that on 5 muM TDZ. Chemical names used: 1-phenyl-3-(1,2,3-thiadiazol-5-yl)urea (thidiazuron, TDZ); 9-(beta-D-ribofuranosyl)-6-(4-hydroxy-3-methyl-but-2-enylamino)purine (zeatin riboside).