940C � Propagation of  Rootstock Tree Production in Greenhouses by Seed, Stem Cuttings and Tissue Culture to Accelerate Budded Tree Production for Outplanting

940C � Propagation of Rootstock Tree Production in Greenhouses by Seed, Stem Cuttings and Tissue Culture to Accelerate Budded Tree Production for Outplanting

Report Date: 01/08/2016
Project: 940C   Year: 2015
Category: Other
Author: Richard Beeson
Sponsor: Citrus Research and Development Foundation

Seed decontamination 1. Develop guidelines for seed propagation that prevent contamination of seed by citrus canker. Surface disinfection of seeds. Objective is to develop economical and easy-to-implement seed cleaning methods for removing Xcc bacteria without loss of seed viability. Progress: Citrus seeds obtained from a local grower were free of any X. citri infection. Therefore citrus seeds were artificially inoculated with X. citri . Seeds were soaked in a freshly prepared X.citri culture (OD 0.002) in a beaker for 1 hour. Contaminate seeds were then placed on sterile filter papers and dried at room temperature for 24 hrs. These were then treated with PPM and Chlorox according to the plan of the project. Seeds were rinsed in sterile water and serial dilutions of the rinsed water were plated for bacterial count. Similarly intact seeds were placed on LB for determining the efficiency of decontamination protocols. Some data has been collected, with the rest being collected from the pictures of LB plates. After completion of data collection, data will be analyzed. The same experiment will be repeated once more. Evaluating the 2006 CHRP decontamination procedure. Progress: These experiments are in progress along with experiments using sodium hypochlorite and PPM for decontamination. Development of rapid and sensitive molecular diagnostic methods for on-site detection of Citrus canker. Progress: Initial standardization experiments are going on. The PI met with our collaborators at UF to obtain additional isolates. Cutting and tissue cultivation On 7 July, 4 schedules of acclimation of tissue cultured citrus rootstocks were applied to C-35, C-54 sour orange, US-812, SW-13 and KC-13 root stocks. Any roots were removed and stems transplanted into a peat:perlite blend in seedling trays. Relative humidity (RH) started at 90% under layers of 30% and 50% shade cloth, overlain with reflective R-3 insulation board (from inside out). RH was decreased step-wise every 2, 3, 4 or 5 days until the end of days at RH = 50%. Covers were similarly removed. Success was very high for most varieties at the fastest acclimatization rate, at over 90%; except sour orange. A second set was started in early October. Cutting propagation began on May 6th and have generally been initiated monthly since. Initially cuttings with 1, 2 or 3 nodes were used. No differences in rooting were noted but multiple nodes required pruning to a single node. Thereafter single node cutting have been used. Different auxin concentrations and carriers were also evaluated. Best results were 4000 and 8000 ppm. Both concentrations have been carried forward. Success has been >90% to date using common rootstocks of Kuharski and C-35.

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