Compatibility of different pollination techniques with microprojectile bombardment of Norway spruce and Scots pine pollen

Transformed pollen could be used in crossings to provide a new approach for producing transgenic seedlings of gymnosperms recalcitrant to tissue culture. The aim of this study was to investigate different pollination techniques in Norway spruce (Picea abies (L.) Karst.) and in Scots pine (Pinus sylvestris L.) using both natural pollen and pollen bombarded with DNA-coated microprojectiles. Cone and seed set after liquid pollinations with natural and bombarded pollen was compared with conventional crossings in which dried natural pollen was used. Liquid pollination proved inferior to conventional crossings. In Norway spruce, the percentages of mature cones and full seeds were lower with liquid pollination than with the conventional method; and in Scots pine, the number of conelets was lower than in pollinations in which dried pollen was used. With bombarded pollen, the results were even lower. To improve possibilities to obtain transgenic progenies, a method was developed that allows the use of bombarded pollen in conventional crossings. This technique is based on dehydration of pollen suspensions and storage at -20 degrees C, which results in a slight decrease in the germination ability of pollen but leads to a high level of the beta-glucuronidase reporter gene expression. This is the first report on transient foreign gene expression in plant material stored in a metabolically inactive form for 1 month before gene expression was assayed.