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BOTANY Characterization of a pollen cDNA library from giant ragweed (Ambrosia trifida L.), and analysis of allergenic pectate lyase. KATHRYNE J. BLAIR*, CHAD M. MANNING, JOANN M. LAU and DAVID L. ROBINSON, Department of Biology, Bellarmine University, Louisville, KY 40205. About 36 million Americans are allergic to pollen released by grasses, trees, and weeds. A major source of this allergenic pollen is giant ragweed (Ambrosia trifida L.), a common weedy plant in the U.S. As of June 2007, there were only 4 nuclear DNA sequences from giant ragweed published in the NCBI GenBank. To examine gene expression in giant ragweed pollen, a cDNA library was constructed from mRNA isolated from dehiscing male flowers. To date, 75 clones have been characterized according to their biological, cellular, and molecular functions using NCBI BLASTX searches. The most common cellular location for these gene products was the nucleus (20%) with 28% appearing to be involved with transcriptional, post-transcriptional, or other DNA/RNA processing events. Examples of gene homologies include a metallothionein, an auxin-response factor, a floral homeotic protein, chitinase, and calmodulin. Several potential allergenic proteins were also isolated, including pectate lyase. Primer walking was performed to determine the entire sequence of the pectate lyase, and PCR primers were designed to isolate the genomic sequence. To look at the expression of pectate lyase in specific tissues, RNA was extracted from multiple tissues, as well as different stages of flower and pollen development. Reverse Transcription-PCR was used on the resulting tissue samples. This research has the potential to reveal much about the biology of this important allergenic plant.
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