Plant Chitinases Research Articles

β-1,3-glucanase and chitinase activities in winter triticales during cold hardening and subsequent infection by Microdochium nivale

AbstractThe accumulation of pathogenesis-related proteins such as β-1,3-glucanases and chitinases was studied in cold induced snow mould resistance in two Polish cultivars of winter triticale, cv. Hewo and cv. Magnat that substantially differ in resistance to Microdochium nivale. The plants were pre-hardened at 12°C for 10 days and hardened at 4°C for 28 days. Subsequently, cold hardened plants were inoculated with fungal mycelium (M. nivale) and incubated at 4°C for 7 days in dark. Cold acclimatisation resulted in suppression of the total glucanase and chitinases activities in the resistant Hewo as well as sensitive Magnat cultivars that possibly coincides with altered metabolism. However, upon infection with M. nivale the chitinases were markedly induced in the cv. Hewo. At the same time, total β-1,3 glucanases activities did not seem to be affected by fungus in any of the tested triticale cultivars. The pattern and/or the activity of chitinases in plants might be indicative for the resistance/susceptibility against M. nivale.

Spore germination of Trichoderma atroviride is inhibited by its LysM protein TAL6

LysM motifs are carbohydrate-binding modules found in prokaryotes and eukaryotes. They have general N-acetylglucosamine binding properties and therefore bind to chitin and related carbohydrates. In plants, plasma-membrane-bound proteins containing LysM motifs are involved in plant defence responses, but also in symbiotic interactions between plants and microorganisms. Filamentous fungi secrete LysM proteins that contain several LysM motifs but no enzymatic modules. In plant pathogenic fungi, for LysM proteins roles in dampening of plant defence responses and protection from plant chitinases were shown. In this study, the carbohydrate-binding specificities and biological function of the LysM protein TAL6 from the plant-beneficial fungus Trichodermaatroviride were investigated. TAL6 contains seven LysM motifs and the sequences of its LysM motifs are very different from other fungal LysM proteins investigated so far. The results showed that TAL6 bound to some forms of polymeric chitin, but not to chito-oligosaccharides. Further, no binding to fungal cell wall preparations was detected. Despite these rather weak carbohydrate-binding properties, a strong inhibitory effect of TAL6 on spore germination was found. TAL6 was shown to specifically inhibit germination of Trichoderma spp., but interestingly not of other fungi. Thus, this protein is involved in self-signalling processes during fungal growth rather than fungal-plant interactions. These data expand the functional repertoire of fungal LysM proteins beyond effectors in plant defence responses and show that fungal LysM proteins are also involved in the self-regulation of fungal growth and development.

Plant growth promoting rhizobacteria (PGPR) and entomopathogenic fungus bioformulation enhance the expression of defense enzymes and pathogenesis-related proteins in groundnut plants against leafminer insect and collar rot pathogen

The bioformulation of Pseudomonas fluorescens (Pf1 and TDK1) and Beauveria bassiana (B2) strains was evaluated individually and in combinations with and without chitin for their efficacy against leafminer insect and collar rot disease and the effect of the interaction between Pseudomonas, Beauveria and groundnut leafminer insect and collar rot pathogen in the expression of defense enzymes and pathogenesis-related proteins (PR-proteins) in groundnut. Among the various bioformulations, B2 + TDK1 + Pf1 (amended with or without chitin) formulation significantly reduced the incidence of leafminer and collar rot disease when compared to untreated control. A significant increase in the enzymatic activity of phenylalanine ammonia-lyase, peroxidase, polyphenol oxidase, chitinase, β-1,3-glucanase, superoxide dismutase, catalase, lipoxygenase, and phenolics in groundnut plants treated with B2 + TDK1 + Pf1 bioformulation (amended with or without chitin) and challenge inoculated with Aproaerema modicella and Sclerotium rolfsii. Native gel electrophoresis also revealed the expression of more isoforms of pathogenesis-related proteins and other defense enzymes viz., polyphenol oxidase and superoxide dismutase in plants treated with B2 + TDK1 + Pf1 mixture challenged with A. modicella and S. rolfsii. The present study reveals that sustained and timely induction and accumulation of these defense enzymes and PR-proteins enhance the resistance in groundnut against leafminer insect and collar rot disease.

Pathogenesis Related Defence Functions of Plant Chitinases and β-1,3-Glucanases

When resistant plants recognize cognate or matching elicitors, intracellular signal transduction pathways are activated that ultimately result in the derepression of a battery of genes called defence response genes. Besides alteration in several biochemical parameters leading to enhanced production of phytoalexins, deposition of callose, lignification etc., the pathogen attack is accompanied by production of several pathogenesis related (PR) proteins. However, only plant chitinases and β-1,3-glucanases have been extensively studied. Pathogenesis-related proteins including hydrolytic enzymes chitinases and β- 1,3-glucanases, have been known to be induced in plants upon infection with various pathogens. This in part is due to the immediate realization that these hydrolytic enzymes could degrade cell walls of certain fungi. These enzymes were shown to be induced in plants upon infection and some purified proteins were observed to have antifungal activity in vitro. Chitinases and glucanases have been purified and characterized from a number of plant sources. Expressions of gene encoding these enzymes have also been quantified from a variety of plant sources. Currently, there is an immense interest in delineating the molecular events from pathogen recognition to the expression of these genes. In an effort to enhance the disease resistance, PR-genes have been used to transform a variety of plant sources. There is considerable evidence showing positive correlation between the expression of PR-genes and disease resistance. The research work being carried out on the role of PR proteins in three important crop species, viz. Sorghum (Sorghum bicolor), rocket salad (Eruca sativa) and moth bean (Vigna aconitifolia) in response to infection by fungal pathogens have been examined.

Potential Application of Chitin Signaling in Engineering Broad-Spectrum Disease Resistance to Fungal and Bacterial Pathogens in Plants

Chitin, a polymer of N-acetylglucosamine, is an important structural component in the cell walls of fungal pathogens. Plant chitinases are capable of degrading this component to directly inhibit infection by fungal pathogens. Plants can further detect the chitin fragments released by chitinases to trigger additional defense responses. Recent studies have identified chitin receptors in different plant species and a number of downstream signaling components. The chitin-mediated signal transduction pathway appears to be a unique and promising target for engineering plants for broad-spectrum resistance to various fungal as well as bacterial pathogens.

Effect of Botrytis cinerea infection and elicitation on ß-1,3-glucanase and chitinase activity in bean leaves and cell cultures

The activity of ß-1,3-glucanase and chitinase in bean plants treated with <i>B. cinerea</i> products or/and infected and in cell cultures after application of fungal products has been studied. <i>Botrytis cinerea</i> infection and culture filtrates, ethanol precipitates, glucan and conidial extract treatment markedly enhanced the activity of both hydrolases. Cell cultures treated with <i>B.cinerea</i> products reacted similarly to intact plants. In plants pretreated with 2-day culture filtrate and conidial extract and then infected, ß-1,3-glucanase and chitinase were induced stronger than after infection without pretreatment.

Chitinase modifying proteins from phylogenetically distinct lineages of Brassica pathogens

Chitinase modifying proteins, cmps, are secreted fungal proteases that truncate specific plant class IV chitinases by cleaving peptide bonds in their amino termini. We recently identified a cmp from the Zea mays (maize) pathogen Fusarium verticillioides and found that it is a member of the fungalysin class of proteases. We also found that Alternaria brassicae, a pathogen of the mustard plant family Brassicaceae, secretes a protease with the same activity. To determine which pathogens of Brassicaceae plants secrete fungalysin cmps, we tested protein extracts from twenty fungi that had been isolated from diseased plants. Each fungal isolate was grown saprotrophically on maize and canola seeds. Secreted fungal proteins were extracted from cultures and incubated with three purified plant chitinases: ChitA and ChitB from maize, and AtchitIV3 from Arabidopsis thaliana. We found that fungalysin cmps were secreted by fungal pathogens distributed among five families in three major Ascomycota classes (Dothideomycetes, Leotiomycetes and Sordariomycetes). Of four fungal species that did not secrete fungalysin cmp activity, three secreted other cmps that truncated maize ChitA and ChitB by cleaving other peptide bonds. AtchitIV3 was only susceptible to truncation by fungalysin cmps. These results show that cmps are commonly secreted by fungal pathogens of Brassicaceae and suggest that interfering with fungalysin cmp activity may improve plant resistance to multiple fungal diseases.

Purification and characterisation of a novel chitinase from persimmon (Diospyros kaki) with antifungal activity

A novel chitinase from the persimmon fruit was isolated, purified and characterised in this report. The Diospyros kaki chitinase (DKC) was found to be a monomer with a molecular mass of 29kDa. It exhibited optimal activity at pH 4.5 with broad pH stability from pH 4.0–9.0. It has an optimal temperature of 60°C and thermostable up to 60°C when incubated for 30min. The internal peptide sequences of DKC showed similarity with other reported plant chitinases. It has the ability to hydrolyse colloidal chitin into chito-oligomers such as chitotriose, chitobiose and into its monomer N-acetylglucosamine. It can be used to degrade chitin waste into useful products such as chito-oligosacchaarides. DKC exhibited antifungal activity towards pathogenic fungus Trichoderma viride. Chitinases with antifungal property can be used as biocontrol agents replacing chemical fungicides.

Transgenic rice lines constitutively co-expressing tlp-D34 and chi11 display enhancement of sheath blight resistance

Transgenic rice (Oryza sativa L. subsp. indica cv. White Ponni) constitutively expressing the rice thaumatin-like protein gene (tlp-D34, PR-5) individually or in combination with the rice chitinase gene (chi11, PR-3) was generated using an Agrobacterium vir helper strain with multiple copies of pTiBo542 virB and virG. Transformation with the tlp-D34 gene alone and tlp-D34 + chi11 genes yielded five and seven single-copy transgenic lines, respectively. Southern blot analysis with two probes, one flanking the right T-DNA border and the second flanking the left T-DNA border, confirmed that all transgenic plants harboured single and complete T-DNA copies. Homozygous transgenic lines were first identified in the T1 generation by Southern blot analysis and were subsequently confirmed by segregation analysis of T2 plants. Accumulation of transcripts encoded by the transgenes was confirmed in T0 plants and homozygous T2 plants by Northern blot analysis. The homozygous T2 plants harbouring tlp-D34 + chi11 genes showed 2.8- to 4.2-fold higher chitinase activity. Western blot analysis revealed the accumulation of thaumatin-like protein and chitinase in the respective transgenic plants. Upon infection with Rhizoctonia solani, the disease index reduced from 100 % in control plants to 65 % in a T3 homozygous transgenic line T4 expressing the tlp-D34 gene alone. In a T2 homozygous transgenic line CT22 co-expressing tlp-D34 and chi11 genes, the disease index reduced to 39 %.

Bacterial and fungal chitinase chiJ orthologs evolve under different selective constraints following horizontal gene transfer

BackgroundCertain bacteria from the genus Streptomyces are currently used as biological control agents against plant pathogenic fungi. Hydrolytic enzymes that degrade fungal cell wall components, such as chitinases, are suggested as one possible mechanism in biocontrol interactions. Adaptive evolution of chitinases are previously reported for plant chitinases involved in defence against fungal pathogens, and in fungal chitinases involved in fungal-fungal interactions. In this study we investigated the molecular evolution of chitinase chiJ in the bacterial genus Streptomyces. In addition, as chiJ orthologs are previously reported in certain fungal species as a result from horizontal gene transfer, we conducted a comparative study of differences in evolutionary patterns between bacterial and fungal taxa.FindingsChiJ contained three sites evolving under strong positive selection and four groups of co-evolving sites. Regions of high amino acid diversity were predicted to be surface-exposed and associated with coil regions that connect certain α-helices and β-strands in the family 18 chitinase TIM barrel structure, but not associated with the catalytic cleft. The comparative study with fungal ChiJ orthologs identified three regions that display signs of type 1 functional divergence, where unique adaptations in the bacterial and fungal taxa are driven by positive selection.ConclusionsThe identified surface-exposed regions of chitinase ChiJ where sequence diversification is driven by positive selection may putatively be related to functional divergence between bacterial and fungal orthologs. These results show that ChiJ orthologs have evolved under different selective constraints following the horizontal gene transfer event.

Sequence analysis and gene expression of putative oil palm chitinase and chitinase-like proteins in response to colonization of Ganoderma boninense and Trichoderma harzianum

Chitinases are glycosyl hydrolases that cleave the β-1,4-glycosidic linkages between N-acetylglucosamine residues in chitin which is a major component of fungal cell wall. Plant chitinases hydrolyze fungal chitin to chitin oligosaccharides that serve as elicitors of plant defense system against fungal pathogens. However, plants synthesize many chitinase isozymes and some of them are not pathogenesis-related. In this study, three full-length cDNA sequences encoding a putative chitinase (EgChit3-1) and two chitinase-like proteins (EgChit1-1 and EgChit5-1) have been cloned from oil palm (Elaeis guineensis) by polymerase chain reaction (PCR). The abundance of these transcripts in the roots and leaves of oil palm seedlings treated with Ganoderma boninense (a fungal pathogen) or Trichoderma harzianum (an avirulent symbiont), and a combination of both fungi at 3, 6 and 12 weeks post infection were profiled by real time quantitative reverse-transcription (qRT)-PCR. Our findings showed that the gene expression of EgChit3-1 increased significantly in the roots of oil palm seedlings treated with either G. boninense or T. harzianum and a combination of both; whereas the gene expression of EgChit1-1 in the treated roots of oil palm seedlings was not significantly higher compared to those of the untreated oil palm roots. The gene expression of EgChit5-1 was only higher in the roots of oil palm seedlings treated with T. harzianum compared to those of the untreated oil palm roots. In addition, the gene expression of EgChit1-1 and EgChit3-1 showed a significantly higher gene expression in the leaf samples of oil palm seedlings treated with either G. boninense or T. harzianum.

The LysM Receptor-Like Kinase LysM RLK1 Is Required to Activate Defense and Abiotic-Stress Responses Induced by Overexpression of Fungal Chitinases in Arabidopsis Plants

Application of crab shell chitin or pentamer chitin oligosaccharide to Arabidopsis seedlings increased tolerance to salinity in wild-type but not in knockout mutants of the LysM Receptor-Like Kinase1 (CERK1/LysM RLK1) gene, known to play a critical role in signaling defense responses induced by exogenous chitin. Arabidopsis plants overexpressing the endochitinase chit36 and hexoaminidase excy1 genes from the fungus Trichoderma asperelleoides T203 showed increased tolerance to salinity, heavy-metal stresses, and Botrytis cinerea infection. Resistant lines, overexpressing fungal chitinases at different levels, were outcrossed to lysm rlk1 mutants. Independent homozygous hybrids lost resistance to biotic and abiotic stresses, despite enhanced chitinase activity. Expression analysis of 270 stress-related genes, including those induced by reactive oxygen species (ROS) and chitin, revealed constant up-regulation (at least twofold) of 10 genes in the chitinase-overexpressing line and an additional 76 salt-induced genes whose expression was not elevated in the lysm rlk1 knockout mutant or the hybrids harboring the mutation. These findings elucidate that chitin-induced signaling mediated by LysM RLK1 receptor is not limited to biotic stress response but also encompasses abiotic-stress signaling and can be conveyed by ectopic expression of chitinases in plants.

Molecular characterization of stress resistance-related chitinase genes of Brassica rapa

Brassica is an important vegetable group worldwide that is impacted by biotic and abiotic stresses. Molecular biology techniques offer the most efficient approach to address these concerns. Inducible plant defense responses include the production of pathogenesis-related (PR) proteins, and chitinases are very important PR proteins. We collected 30 chitinase like genes, three from our full-length cDNA library of Brassica rapa cv. Osome and 27 from Brassica databases. Sequence analysis and comparison study confirmed that they were all class I-V and VII chitinase genes. These genes also showed a high degree of homology with other biotic stress resistance-related plant chitinases. An organ-specific expression of these genes was observed and among these, seven genes showed significant responses after infection with Fusarium oxysporum f.sp. conglutinans in cabbage and sixteen genes showed responsive expression after abiotic stress treatments in Chinese cabbage. BrCLP1, 8, 10, 17 and 18 responded commonly after biotic and abiotic stress treatments indicating their higher potentials. Taken together, the results presented herein suggest that these chitinase genes may be useful resources in the development of stress resistant Brassica.

Brittle culm15 encodes a membrane-associated chitinase-like protein required for cellulose biosynthesis in rice.

Plant chitinases, a class of glycosyl hydrolases, participate in various aspects of normal plant growth and development, including cell wall metabolism and disease resistance. The rice (Oryza sativa) genome encodes 37 putative chitinases and chitinase-like proteins. However, none of them has been characterized at the genetic level. In this study, we report the isolation of a brittle culm mutant, bc15, and the map-based cloning of the BC15/OsCTL1 (for chitinase-like1) gene affected in the mutant. The gene encodes the rice chitinase-like protein BC15/OsCTL1. Mutation of BC15/OsCTL1 causes reduced cellulose content and mechanical strength without obvious alterations in plant growth. Bioinformatic analyses indicated that BC15/OsCTL1 is a class II chitinase-like protein that is devoid of both an amino-terminal cysteine-rich domain and the chitinase activity motif H-E-T-T but possesses an amino-terminal transmembrane domain. Biochemical assays demonstrated that BC15/OsCTL1 is a Golgi-localized type II membrane protein that lacks classical chitinase activity. Quantitative real-time polymerase chain reaction and β-glucuronidase activity analyses indicated that BC15/OsCTL1 is ubiquitously expressed. Investigation of the global expression profile of wild-type and bc15 plants, using Illumina RNA sequencing, further suggested a possible mechanism by which BC15/OsCTL1 mediates cellulose biosynthesis and cell wall remodeling. Our findings provide genetic evidence of a role for plant chitinases in cellulose biosynthesis in rice, which appears to differ from their roles as revealed by analysis of Arabidopsis (Arabidopsis thaliana).

Tree chitinases - stress- and developmental-driven gene regulation

Summary In recent years, a considerable number of studies have harnessed the power of genomics to decipher the role of pathogenesis-related (PR) proteins in plant defence against various biotic and abiotic stresses. Chitinases are PR antifungal proteins expressed constitutively at low levels in plants and induced during biotic pressures and are demonstrated to be involved in the plant defence responses. Remarkable induction of chitinase enzymes by various abiotic agents (salicylic acid, jasmonic acid, ethylene and ozone) and biotic components (pathogens, insect pest, fungal cell wall components and oligosaccharides) is well demonstrated in plants. Several reviews on plant chitinase expression during host–pathogen interaction are available for annual species, whilst reports of their expression in tree species are limited to a few woody perennials: Populus, Pinus, Picea, Eucalyptus, Castanea and Pseudotsuga. The aim of this paper is to review the induction of chitinase during various stresses and developmental processes in forest tree species.

Plant Chitinases: Genetic Diversity and Physiological Roles

Chitinase proteins are widely distributed across diverse biological systems. Chitinases hydrolyze chitin, chitosan, lipochitooligosaccharides, peptidoglycan, arabinogalactan and glycoproteins containing N-acetylglucosamine. Analyses of genome-wide sequence and microarray expression profilings show that chitinase genes are represented by large families and the individual member genes are expressed in diverse conditions. Chitinase proteins are members in the group of the pathogenesis-related proteins that are strongly induced when host plant cells are challenged by pathogen stress and thus chitinases constitute an important arsenal of plants against fungal pathogens. Transgenic plants have been produced that overexpress chitinases alone or in conjunction with other defense-related proteins. The phenotype analyses of such plants have shown enhanced disease resistance in large number of cases. Apart from defense against pathogen stress, chitinases are implicated in relationships between plant cells and fungi (e.g., mycorrhizae associations) and bacteria (e.g., legume/Rhizobium associations). Chitinases are also involved in plant abiotic stress responses as noted for osmotic, salt, cold, wounding and heavy metal stresses. Chitinases play a role in developmental aspects of plants too (i.e., regulation of plant embryogenesis process). A detailed account of the genetic diversity and functional aspects of plant chitinases is presented in this review.

Plant chitinase III Ziz m 1 stimulates multiple cytokines, most predominantly interleukin-13, from peripheral blood mononuclear cells of latex-fruit allergic patients

BackgroundIndian jujube is a fruit abundantly cultivated in Taiwan. Its major allergen in latex-fruit syndrome is Ziz m 1 of the chitinase III family. The Ziz m 1 Pichia (rZiz m 1-P) has chitinase activity but not Ziz m 1 E. coli (rZiz m 1-E). ObjectiveThis study examined whether plant chitinase III, using rZiz m 1-P and rZiz m 1-E, can stimulate allergic inflammation similar to that of mammalian chitinases. MethodsFive patients allergic to latex–Indian jujube and five nonallergic controls were evaluated. Their peripheral blood mononuclear cells (PBMC) were cultured with rZiz m 1-E or rZiz m 1-P and pulsed with phorbol 12-myristate 13-acetate. Eleven cytokines were measured by FlowCytomix human Th1/Th2 plex kit and interleukin (IL)-13 by sandwich enzyme-linked immunosorbent assay (ELISA). ResultsInterleukin-13 significantly increased in rZiz m 1-P stimulated PBMC of allergic subjects but was undetectable when stimulated with rZiz m 1-E. The stimulation index significantly increased in IL-13 (380.6 ± 77.33 vs 13.70 ± 6.92), IL-5 (6.70 ± 0.59 vs 0.70 ± 0.37), IL-1β (32.70 ± 0.83 vs 2.10 ± 1.29), and tumor necrosis factor beta (TNF-β) (17.10 ± 2.66 vs 1.50 ± 0.66) between allergic and nonallergic subjects after rZiz m 1-P stimulation. There was no difference in terms of IL-2, IFN-γ, IL-8, and TNF-α production. ConclusionsThe biological function of chitinase activity is required for Ziz m 1 to induce a Th2-specific immune response. This is the first report on PBMC responses of latex-fruit syndrome subjects toward an active exogenous plant class III chitinase that can stimulate multiple cytokines, especially IL-13, from allergic subjects. This implies the role of cross-reactive food allergens in propagating allergic inflammation among allergic subjects.

Effect of phytoplasma infection on the activity of peroxidase, β-1,3 glucanase and chitinase in corn plants

In the present work we studied the effect of inoculating corn plants with the maize bushy stunt phytoplasma on the activity of the enzymes peroxidase, β-1,3 glucanase and chitinase. The experiments were carried out inside a greenhouse. Plants of a resistant and a susceptible corn hybrid were inoculated by using infective Dalbulus maidis leafhoppers 10 days after sowing. When symptoms started to appear, leaf samples were collected at different periods to quantify enzyme activity. The results showed an increase in the activity of the three enzymes in inoculated plants of both hybrids. In general, the values observed for the level of the different enzymes were higher in the susceptible hybrid when compared to the resistant one. Thus, the increases in peroxidase, β-1,3 glucanase and chitinase levels in inoculated plants are evidence of changes in the host metabolism caused by the phytoplasma. On the other hand, since the increases could not be correlated with plant resistance further studies are needed to explain such changes.

Biological control of sheath blight disease of rice by siderophore producing rhizobacterial strains and their role in efficient mobilisation of micronutrients from soil

Four siderophore producing rhizobacterial strains Bacillus cereus Z2-7, Enterobacter sp. B41 strain SPR7, Aeromonas hydrophila BPS10 and Enterobacter sp. BPS12 isolated from rice grown in saline soils were examined for induction of chitinases, glucanases and peroxideses (POX) in rice plants against sheath blight pathogen R. solani and mobilization of micronutrients from rice rhizosphere to straw and grain of rice plant. Atomic absorption studies indicated that rice plants preinoculated with Bacillus cereus Z2-7 have an increased uptake of zinc, copper and iron as compared to uninoculated control plants. High content of copper and iron were present in straw while high quantity of zinc was found in grains. Straw of rice plants inoculated with Enterobacter sp. B41 strain SPR7 contained highest Manganese content as compared to other treatments and uninoculated control plants. Gene expression analysis revealed enhanced levels of chitinase, peroxidase and glucanase expression in rice plants preinoclated with bacterial strains Z2-7 and SPR7 after 7 and 52 days of challenge inoculation with R. solani. Improved uptake of micronutrients and induction of defence enzymes in these plants resulted in an increase in yield of these plants as compared to rest of the treatments.

Identification and expression analysis of chitinase genes related to biotic stress resistance in Brassica

Brassica is a very important vegetable group because of its contribution to human nutrition and consequent economic benefits. However, biotic stress is a major concern for these crops and molecular biology techniques offer the most efficient of approaches to address this concern. Chitinase is an important biotic stress resistance-related gene. We identified three genes designated as Brassica chitinase like protein (BrCLP1), BrCLP2 and BrCLP3 from a full-length cDNA library of Brassica rapa cv. Osome. Sequence analysis of these genes confirmed that BrCLP1 was a class IV chitinase, and BrCLP2 and BrCLP3 were class VII chitinases. Also, these genes showed a high degree of homology with other biotic stress resistance-related plant chitinases. In expression analysis, organ-specific expression of all three genes was high except BrCLP1 in all the organs tested and BrCLP2 showed the highest expression compared to the other genes in flower buds. All these genes also showed expression during all developmental growth stages of Chinese cabbage. In addition, BrCLP1 was up-regulated with certain time of infection by Pectobacterium carotovorum subsp. carotovorum in Chinese cabbage plants during microarray expression analysis. On the other hand, expression of BrCLP2 and BrCLP3 were increased after 6h post inoculation (hpi) but decreased from 12hpi. All these data suggest that these three chitinase genes may be involved in plant resistance against biotic stresses.