Studies on factors involved in transfer of foreign gene by pipetting Agrobacterium into spikelets of wheat.

摘要

To determine the various factors involved in gene transfer, studies were conducted on wheat cv. UP 2338 by pipetting 5μl Agrobacterium suspension into florets using indirect pollen system. Pollen germination, Agrobacterium staining and grain screening tests were conducted. Results showed that 20 minutes after pollination was the best time for Agrobacterium (LBA4404 containing binary vector pBI121) inoculation. After 20 minutes of pollination, pollen grains showed germination with an average germination of 45.4%. The emergence of pollen tubes in the hairy stigma and the bifurcated style was seen after 30 minutes of pollination. Gram negative staining for Agrobacterium resulted in a pink colour 20 minutes after Agrobacterium inoculation. At later stages, some parts of the pistil and pollen turned pink. Seeds obtained after Agrobacterium inoculation into the florets showed a 5.5% putative transformation frequency when screened against 500 mg/l kanamycin. Based on phenotypic observations, it can beconcluded that it is possible to transfer foreign gene via pollen through Agrobacterium.