Chitinase has been suggested to play a role in defense mechanisms. In this study, the activity and expression of chitinase in muskmelon seeds were investigated. Multiple chitinase isoforms were detected in muskmelon seeds from early development through radicle emergence. One acidic and three basic chitinase isoforms were detected in developing seeds at 40 days after anthesis (DAA). Both acidic and basic chitinase isoforms were detected in endosperm tissue during seed imbibition and after radicle emergence. Basic chitinase isoforms, but not acidic isoforms, were detected in embryo tissue. Basic chitinase isoforms were also detected in the embryonic axis or radicle tissue. Taken together, these observations indicate that chitinases are regulated developmentally and in a tissue-specific manner in muskmelon seeds. Therefore the potential function of chitinases in muskmelon seeds is discussed. Two complete cDNAs, Cmchi1 and Cmchi2, and a partial genomic clone of Cmchi2 have been isolated from muskmelon seeds. Cmchi2 gene has two introns in the coding region while Cmchi1 is intronless. Cmchi1 cDNA encodes a class III chitinase while Cmchi2 cDNA encodes a class II chitinase. Cmchi1 and Cmchi2 proteins might be targeted to secretory pathways because they possess signal peptides. Southern blotting suggested that there is at least one additional gene similar to Cmchi1 in the muskmelon seed genome, while there is only one copy of Cmchi2. Northern blotting analysis showed that both Cmchi1 and Cmchi2 are expressed in the radicle tissue at the time of radicle emergence. This indicates that the expression is regulated developmentally and in a tissue-specific manner. Salicylic acid (SA) and benzothiadiazole (BTH) stimulated the expression of Cmchi1 but not Cmchi2 in seeds after radicle emergence, indicating that SA might be involved in inducing the expression of Cmchi1, while a different signal might be involved in triggering the expression of Cmchi2. The protein encoded by Cmchi1cDNA was expressed in E.coli. It did not show any enzymatic activity. Western blotting using an antibody raised against the class III chitinase protein in cucumber was inconclusive, as this antibody recognized the purified Cmchi1 fusion protein and other unknown proteins isolated from the embryonic axis or the radicle tissue. / Ph. D.

Respiration rates of developing muskmelon (Cucumis melo L.) seeds were determined polarographically using a Clark-type O₂ electrode (Hansatech LD2). Seeds were obtained from fruits harvested 20, 30, 40, and 50 days after anthesis (DAA). Respiration (O₂ uptake) was measured for fresh intact seeds and fresh dissected seeds. The respiration rate of intact seeds declined from a maximum of 2.28 μmol O₂/min/g DWT at 20 DAA to a minimum of 0.16 μmol O₂/min/g DWT at 50 DAA. Dissecting intact seeds into embryo, testae, and perisperm tissues increased the respiration rate of 20 DAA seeds to 3.12 μmol O₂/min/g DWT but had no effect on more mature seeds. Respiration rate was highly correlated with seed relative growth rate and water content. Respiration rate was not consistently changed after incubation in water. This indicates that respiration rate is not directly controlled by subtle variations in water content. Rather, seed respiration rate is directly linked with turgor-driven, expansive growth and relative growth rate. Fifty-DAA seeds from dry storage were imbibed on water saturated blotters, and respiration rates of whole seeds, decoated seeds, and embryos were compared. Respiration during imbibition was not significantly inhibited by the testae or perisperm tissue. In addition, 50-DAA dried, imbibed seeds were subjected to reduced O₂ concentrations ranging from 3.5 kPa partial pressure O₂ (pO₂) to 21 kPa pO₂. Respiration was not limited by O₂ until pO₂ was reduced to approximately 5 kPa, indicating a high affinity for O₂. Gas chromatography revealed that pO₂ in the seed cavity of muskmelon fruits ranged from 12.5 to 8 kPa. Fifty-DAA seeds from dry storage were imbibed on polyethylene glycol (PEG), mannitol, or NaCl ranging from -0.5 to -2.5 MPa water-potential or on abscisic acid (ABA) solutions ranging in concentration from 10 to 50 μM. Respiration and solution water-potential were measured at 10-hr intervals. At 10 hr of imbibition, each type of osmoticum and ABA stimulated respiration to values greater than for seeds imbibed in pure water. Beyond 10 hr, respiration rates were variable. / Master of Science

Call number: LD2668 .T4 1984 B425 / Master of Science