( ns have been demonstrated using the PDS...)Optimization of Biolistic Transformation Using the Helium-Driven PDS-1000/He System

ns have been demonstrated using the PDS- 1000/He instrument. Transformation efficiencies can be maximized by optimizing the microparticle size, target distance, and helium pressure. In both systems studied here, the first two parameters are the most important. Little difference in results was found using helium pressures between 900 and 1,550 psi. This report also shows that these conditions may be optimized in one or two experiments; because of shot-to-shot variability, it is important to perform at least three to five replicates for each set of conditions. The optimum parameters found for the yeast and cauliflower systems assayed here are summarized in Table 6. When optimizing other systems, we suggest to begin by using conditions given in Table 7, then examine in order: target distance, microparticle size and type, and helium pressure.

Biolistic is a registered trademark of E. I. du Pont de Nemours & Co. The Biolistic technology is licensed to Bio-Rad Laboratories.

Appendices
The procedures described in Appendices 1 and 2 were developed by Sanford, et al. (1992).

Appendix 1. Microcarrier preparation
For 120 bombardments using 500 g per bombardment

1. In a 1.5 ml microfuge tube, weigh out 60 mg of microparticles.

2. Add 1 ml of 70% ethanol, freshly prepared.

3. Vortex on a platform vortexer for 3-5 minutes.

4. Incubate for 15 minutes.

5. Pellet the microparticles by spinning for 5 seconds in a microfuge.

6. Remove the liquid and discard.

7. Repeat the following steps three times:
a. Add 1 ml of sterile water.
b. Vortex for 1 minute.
c. Allow the particles to settle for 1 minute.

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