| Scope and method of study. This research involved the characterization and functional analysis of two extensins (extensins # 2 and # 3), and an LRR-extensin (# 65631). 5' RACE experiment was done to determine the 5' ends of the LRR-extensin, and the genomic clone was isolated after screening the genomic library of cotton with extensin specific probes. Tissue expression analysis was done by northern blots to find out the expression pattern of the extensins and LRR-extensin in the different tissues of cotton. Infection and wounding studies were done on cotton cotyledons to see if the extensins and LRR-extensins were defense-related genes. The LRR domain of the LRR extensin was expressed in a prokaryotic expression vector, and polyclonal antibodies were synthesized to the LRR domain. Western blot analysis was done using these antibodies to detect the molecular weight of the LRR-extensin in plant tissues.; Findings and conclusions. The LRR-extensin is a 2.46 kb transcript as demonstrated by RACE, Northern blot and genomic library screening. It is expressed predominantly in stem, cotyledons, callus, and leaves. Infection studies show that the expression of this gene is not induced at 48 hours after infiltration with Xanthomonas campestris pv. vesicatoria. Cotton LRR-extensin is not induced in cotyledons 48 hours after wounding. Western blot results with anti-LRR polyclonal antibodies, recognized a protein of 300 kD in callus cultures. Clones for Extensin # 2 and # 3 are 1 kb and 700 bp long, respectively, and both are missing their 5' ends. They are mainly expressed in root, stem and callus. Infection studies show that both the extensins are induced at 48 hours after infection, and wounding studies show that both the genes are expressed at 12 hours after wounding the cotyledons. From these results, one can conclude that the extensins are defense related genes and the LRR-extensin is not involved in plant defense. |
