As part of a program to develop transgenic peach (Prunus persica L. Batsch) cultivars with resistance to Prunus necrotic ringspot virus (PNRSV), we are testing a system for measuring virus in peach shoot cultures. Micrografting in vitro is used for inoculation and slot-blot hybridization, with a digoxigenin (DIG)-labeled cRNA probe complementary to the 5′ open reading frame (ORF) of PNRSV RNA 3, for detection. In this study, we investigated whether infected shoots maintain virus infection over long periods of culture at 4 °C and if PNRSV-infected `Suncrest' shoot cultures can serve as graft bases to transmit virus equally well into cultivars Nemaguard, Springcrest, and Suncrest. The results of RNA hybridization analysis showed that virus was present in extracts of leaf samples from 2-year-old PNRSV-infected `Suncrest' shoots that had been subjected to varying lengths of incubation at 4 °C in the dark, suggesting that infected shoots can be maintained for repeated use. Rates of graft success were higher in heterografts between `Suncrest' bases and tips of `Springcrest' or `Nemaguard' than in autografts between `Suncrest' and `Suncrest', and there was equal efficacy of graft inoculation from `Suncrest' into these three cultivars.
K. Heuss, Q. Liu, F.A. Hammerschlag, and R.W. Hammond
Z. Wang, M.C. Acock, Q. Liu, and B. Acock
Flowering time, growth, and opium gum yield from five seed sources (T, L, B1, B2, B3) of opium poppy (Papaver somniferum L.) collected from different latitudes in three Southeast Asian countries were determined. Plants were grown in six growth chambers at a 11-, 12-, 13-, 14-, 15-, or 16-hour photoperiod with a 12-hour, 25/20 °C thermoperiod. Flower initiation was observed under a dissecting microscope (40×) to determine if time to floral initiation was identical for all accessions across a wide range of photoperiods. The main capsule was lanced for opium gum at 10, 13, and 16 days after flowering (DAF). Plants were harvested at 21 DAF for plant height, leaf area, and organ dry-weight determinations. In a 16-hour photoperiod, flower initiation was observed 10 days after emergence (DAE) for B1 vs. 8 DAE for the other four accessions. Flowering time was affected most by photoperiod in B1 and least in B2. Flowering times for B3, L, and T were similar across the range of photoperiods. B2, B3, and L had the highest gum yields per capsule; even though B1 had the greatest total plant biomass, it produced the lowest gum yield. There was no difference among accessions in the average ratio of gum: individual capsule volume. For the ratio of gum: capsule dry weight, only the difference between T and B1 was significant. Capsule size did affect these ratios slightly. T had a larger gum: volume ratio for larger capsules, and B3 had a smaller gum: dry-weight ratio for heavier capsules. Flowering time varied up to 40%, capsule dry weight up to 41%, and opium gum yield up to 71% for the five accessions across all treatments. No relationship was found between flowering time and the latitude where the seed sources were collected. Time to flower initiation could not be used to predict time to anthesis because floral development rates varied significantly among accessions and photoperiods. Capsule volume and dry weight were useful in estimating gum yield.
Q. Liu, S. Salih, J. Ingersoll, R. Meng, L. Owens, and F. Hammerschlag
Transgenic `Royal Gala' apple (Malus × domestica Borkh.) shoots were obtained by Agrobacterium-mediated gene transfer using the plasmid binary vector pGV-osm-AC with a T-DNA encoding a chimeric gene consisting of a secretory sequence from barley-amylase joined to the modified cecropin MB39 coding sequence. Shoots were placed under the control of a wound-inducible, osmotin promoter from tobacco. The integration of the cecropin MB39 gene into apple was confirmed by Southern blot analysis. The transformation efficiency was 1.5% when internodes from etiolated shoots were used as explants and 2% when leaf explants were used. Both non- and transgenic tetraploid plants were produced by treatment of leaf explants with colchicine at 25 mg·L-1, and polyploidy was confirmed by flow cytometry. Of the diploid transgenics, three of seven were significantly more resistant to Erwinia amylovora than the non-transgenic `Royal Gala' control. Also, in one instance, a tetraploid transgenic was significantly more resistant than the diploid shoot from which it was derived.