Comparative proteomic analysis of cauliflower under high temperature and flooding stresses.

High-temperature and waterlogging are major abiotic stresses that affect the yield and quality of cauliflower. Cauliflower cultivars 'H41' and 'H69' are tolerant to high temperature and flooding, respectively; however, 'H71' is sensitive to both stresses. The objectives of this study were to identify the proteins that were differentially regulated and the physiological changes that occurred during different time periods in 'H41', 'H69', and 'H71' when responding to treatments of flooding, 40 °C, and both stresses combined. Changes in the leaf proteome were analyzed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) and identified by Mascot peptide mass fingerprint (PMF) and database searching. Stress treatments caused significant reductions in electrolyte leakage, chlorophyll fluorescence Fv/Fm, chlorophyll content, and water potential as stress times were prolonged. By the comparative proteomic analysis, 85 protein peaks that were differentially expressed in response to combination treatments at 0, 6, and 24 h, 69 (33 in 'H41', 29 in 'H69', and 9 in 'H71') were identified, of which were cultivar specific. Differentially regulated proteins predominantly functioned in photosynthesis and to a lesser extent in energy metabolism, cellular homeostasis, transcription and translation, signal transduction, and protein biosynthesis. This is the first report that utilizes proteomics to discover changes in the protein expression profile of cauliflower in response to heat and flooding.

Vaccine-associated rhabdomyosarcoma with spinal epidural invasion and pulmonary metastasis in a cat.

A 7-year-old, female, domestic medium-haired cat had a recurrent deep dermal mass in the interscapular region after initial surgical removal 3 months earlier. The cat had received a killed rabies vaccine and a five-in-one vaccine in the same area about 2 months prior to the first surgery. The relapsed mass was diagnosed as vaccine-associated sarcoma. The cat was euthanized 2 months later because of hind limb paralysis. At necropsy, multiple, poorly demarcated, nodular masses were seen in the muscles around the shoulders, neck, and thoracic vertebrae. Pulmonary metastasis and spinal epidural invasion at T1-T3 with regional cord compression and malacia were observed. Microscopically, the masses consisted of interwoven bundles of spindle cells with prominent multinucleated giant cell formation. The neoplastic cells stained strongly positive for myoglobin, and moderately but variably positive for vimentin, desmin, and alpha- smooth muscle actin. Phosphotungstic acid-hematoxylin staining revealed cytoplasmic striations in scattered tumor cells. The tumor was considered a vaccine-associated rhabdomyosarcoma.

The N-terminal signal sequence and the last 98 amino acids are not essential for the secretion of Bacillus sp. TS-23 alpha-amylase in Escherichia coli.

A truncated Bacillus sp. TS-23 alpha-amylase gene lacking 96 and 294 bp at its 5' and 3' end respectively was prepared by polymerase chain reaction and cloned into Escherichia coli expression vector, pQE-30, under the control of T5 promoter. SDS-PAGE and activity staining analyses showed that the His6-tagged amylase had a molecular mass of approximately 54 kDa. Isopropyl-beta-d-thiogalactopyranoside (IPTG) induction of E. coli M15 cells bearing the recombinant plasmid resulted in the extracellular production of active amylase. Western blot analysis also revealed that the truncated amylase was present in the periplasmic space and culture medium.

Purification and properties of chitinase from cabbage.

Chitinase has been purified from the extract of cabbage through successive steps of ammonium sulfate fractionation, chromatofocusing and Sephadex G-75 gel filtration. By these steps, the purity of the enzyme increased by 93.3 fold and the recovery of the enzyme activity was 20%. The purified enzyme had an optimal pH of 5.0, an optimal temperature between 40 to 50 degrees C and a Km of 76 microM for hydrolysis of ethylene glycol chitin. The molecular weight of the enzyme determined from filtration through Sephadex G-75 was 30,000 daltons. Heavy metal ions, Hg2+ (0.5 mM) and Ag+(2.5 mM) significantly inhibited the activity of the enzyme. NBSI1 (1.0 mM), DNFB (0.5 mM) and PMSF (0.5 mM) completely inhibited the activity of the enzyme. The enzyme also showed muramidase activity for hydrolysis of Micrococcus lysodeikticus cell wall. The presence of chitinase in cabbage may function as a defense enzyme against potential pathogens.