Quantitative analysis of chitinase gene expression in chickpea (original) (raw)
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International Journal of Molecular Sciences
Chitinases, a subgroup of pathogenesis-related proteins, are responsible for catalyzing the hydrolysis of chitin. Accumulating reports indicate that chitinases play a key role in plant defense against chitin-containing pathogens and are therefore good targets for defense response studies. Here, we undertook an integrated bioinformatic and expression analysis of the cucumber chitinases gene family to identify its role in defense against Fusarium oxysporum f. sp. cucumerinum. A total of 28 putative chitinase genes were identified in the cucumber genome and classified into five classes based on their conserved catalytic and binding domains. The expansion of the chitinase gene family was due mainly to tandem duplication events. The expression pattern of chitinase genes was organ-specific and 14 genes were differentially expressed in response to F. oxysporum challenge of fusarium wilt-susceptible and resistant lines. Furthermore, a class I chitinase, CsChi23, was constitutively expressed...
Current View on Chitinase for Plant Defence
Plant pathogen is serious problem worldwide amongst the crop cultivars. Large number of plants suffers from range of infectious diseases caused by different pathogens, amongst them fungi are responsible for majority of infectious plant diseases that limiting the crop yield and impact the post-harvest quality of food. So it is important to protect the plants from fungal infection. Substantial approaches have been done with fungicides or heavy metals and by conventional breeding for combating the fungal diseases but so far no conclusive solution has been developed. Genetic engineering paves a new way to protect the plants from harmful fungal afflictions by introducing genes encoding chitinase enzyme that degrade the chitin, which are the key component of fungal cell wall.
Assessment of Differential Expression of Chitinase Genes Using RT-PCR in Chickpea
Biotechnology & Biotechnological Equipment, 2001
In this study, we have found differences in chitinase gene expression between sensitive (Red chickpea) and resistant (Flip 9 I-4C) chickpea (Cicer arietinwn L.) genotypes to Ascochyta blight using Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) technique. Total RNAs extracted from leaves of sensitive and resistant genotypes were used for RT-PCR analysis with chitinase gene :;pecific primer (chi) and oligo dT 15 primer. Expression of chitinase genes was found to be different in the leaves of sensitive and resistant genotypes using this technique.
… and molecular plant …, 2006
Differentially expressed genes in chickpea, (Cicer arietinum L.) during root infection by Fusarium oxysporum f sp. ciceri Race1, were identified using cDNA-RAPD and cDNA-AFLP approaches. The former employed decamer primers on cDNA template and revealed nine differentially expressed transcripts in the resistant-infected chickpea variety. Among the 2000 transcript-derived fragments (TDFs) screened by cDNA-AFLP, 273 were differentially expressed in chickpea roots during Fusarium infection. Only 13.65% of the TDFs were differentially regulated during pathogen challenge, while the other 86% were expressed non-differentially during the process of pathogen infection in chickpea roots. Nineteen TDFs, which expressed differentially in the resistant-infected chickpea variety were cloned and sequenced. Two of these TDFs were similar to transcription factors like WRKY proteins and 14-3-3 proteins, while three TDFs represented the NBS-LRR-type gene sequences. Two TDFs had sequence identity to genes known to have function in defense. The RAPD TDF CaFRi60 showed sequence identity to gamma-glutamyl-cysteine synthetase. Among the TDFs examined by cDNA-AFLP, 19 were confirmed by Reverse Northern blot to be differentially expressed. The data confirms the effectiveness of the cDNA-AFLP technique in detecting differentially expressed genes during pathogenesis. r
The plant pathology journal, 2015
This is the first study reporting the evaluation of transgenic lines of tomato harboring rice chitinase (RCG3) gene for resistance to two important fungal pathogens Fusarium oxysporum f. sp. lycopersici (Fol) causing fusarium wilt and Alternaria solani causing early blight (EB). In this study, three transgenic lines TL1, TL2 and TL3 of tomato Solanum lycopersicum Mill. cv. Riogrande genetically engineered with rice chitinase (RCG 3) gene and their R1 progeny was tested for resistance to Fol by root dip method and A. solani by detached leaf assay. All the R0 transgenic lines were highly resistant to these fungal pathogens compared to non-transgenic control plants. The pattern of segregation of three independent transformant for Fol and A. solani was also studied. Mendelian segregation was observed in transgenic lines 2 and 3 while it was not observed in transgenic line 1. It was concluded that introduction of chitinase gene in susceptible cultivar of tomato not only enhanced the resi...
Effect of chitinase antisense RNA expression on disease susceptibility of Arabidopsis plants
Plant Molecular Biology, 1994
Chitinases accumulate in higher plants upon pathogen attack are capable of hydrolyzing chitin-containing fungal cell walls and are thus implicated as part of the plant defense response to fungal pathogens. To evaluate the relative role of the predominate chitinase (class I, basic enzyme) ofArabidopsis thaliana in disease resistance, transgenic Arabidopsis plants were generated that expressed antisense RNA to the class I chitinase. Young plants or young leaves of some plants expressing antisense RNA had < 10~o of the chitinase levels of control plants. In the oldest leaves of these antisense plants, chitinase levels rose to 37-90~o of the chitinase levels relative to vector control plants, most likely because of accumulation and storage of the enzyme in vacuoles. The rate of infection by the fungal pathogen Botrytis cinerea was measured in detached leaves containing 7-15 ~o of the chitinase levels of control plants prior to inoculation. Antisense RNA was not effective in suppressing induced chitinase expression upon infection as chitinase levels increased in antisense leaves to 47~o of levels in control leaves within 24 hours after inoculation. Leaves from antisense plants became diseased at a slightly faster rate than leaves from control plants, but differences were not significant due to high variability. Although the tendency to increased susceptibility in antisense plants suggests that chitinases may slow the growth of invading fungal pathogens, the overall contribution of chitinase to the inducible defense reponses in Arabidopsis remains unclear.
BMC biotechnology, 2024
Fusarium head blight (FHB) is a devastating fungal disease affecting different cereals, particularly wheat, and poses a serious threat to global wheat production. Chitinases and β-glucanases are two important proteins involved in lysing fungal cell walls by targeting essential macromolecular components, including chitin and β-glucan micro fibrils. In our experiment, a transgenic wheat (Triticum aestivum) was generated by introducing chitinase and glucanase genes using Biolistic technique and Recombinant pBI121 plasmid (pBI-ChiGlu (-)). This plasmid contained chitinase and glucanase genes as well as nptII gene as a selectable marker. The expression of chitinase and glucanase was individually controlled by CaMV35S promoter and Nos terminator. Immature embryo explants from five Iranian cultivars (Arta, Moghan, Sisun, Gascogen and A-Line) were excised from seeds and cultured on callus induction medium to generate embryonic calluses. Embryogenic calluses with light cream color and brittle texture were selected and bombarded using gold nanoparticles coated with the recombinant pBI-ChiGlu plasmid. Bombarded calluses initially were transferred to selective callus induction medium, and later, they were transfferd to selective regeneration medium. The selective agent was kanamycin at a concentration of 25 mg/l in both media. Among five studied cultivars, A-Line showed the highest transformation percentage (4.8%), followed by the Sisun, Gascogen and Arta in descending order. PCR and Southern blot analysis confirmed the integration of genes into the genome of wheat cultivars. Furthermore, in an in-vitro assay, the growth of Fusarium graminearum was significantly inhibited by using 200 μg of leaf protein extract from transgenic plants. According to our results, the transgenic plants (T 1) showed the resistance against Fusarium when were compared to the non-transgenic plants. All transgenic plants showed normal fertility and no abnormal response was observed in their growth and development.
Journal of Plant Physiology, 2006
Fusarium oxysporum f. sp. melonis is a highly specialized fungus that attacks the root system of melon (Cucumis melo L.). In this work the presence of a class III chitinase was examined by immunological techniques in the root and stem base of a susceptible (cv. Galia) and a resistant (cv. Bredor) melon during the infection process. By immunolocalization it was not possible to detect the constitutive presence of class III chitinase in any of the cultivars. However, the immunolabelling appeared in the root tissues of both cultivars as a consequence of wounding and of infection by F. oxysporum f. sp. melonis. Distinct patterns of chitinase detection were observed in the roots of the two cultivars as the infection progressed. Furthermore, by western blotting distinct class III chitinase isoforms were detected, which responded differently to the F. oxysporum f. sp. melonis infection. Our results strongly indicate that a relationship exists between class III chitinase and melon resistance to Fusarium infection, and that the resistance is associated with certain isoforms of this enzyme.
TAG Theoretical and Applied Genetics, 2001
Chitinases and β-1,3-glucanases are important components of plant defense in response to attack by pathogens. To identify specific chitinases and β-1,3-glucanases, we constructed a cDNA library using mRNA from wheat spikelets inoculated with conidia of Fusarium graminearum. Two chitinase and two β-1,3-glucanase clones were isolated using a rice chitinase Ia gene and barley cDNA clones for chitinase II and β-1,3-glucanase as probes. Sequence analysis showed that the cDNA clone SM194 encodes an acidic isoform of class-VII chitinase, the cDNA clone SM383 encodes a class-IV chitinase and the cDNA clones SM289 and SM638 encode two different acidic isoforms of β-1,3-glucanases. Nulli-tetrasomic analysis indicated that SM194 and SM383 were located on all of the group-2 chromosomes of wheat. Genetic mapping showed that at least three copies of class-IV and/or class-VII chitinase genes were clustered on the long arm of chromosome 2D of Aegilops tauschii and that they mapped genetically close to the centromere. SM289 and SM638 were located on all of the group 3 chromosomes of wheat by nulli-tetrasomic analysis, and to the β-1,3-glucanase clusters in the 3BL and 3DL chromosome arms of wheat by genetic mapping. Northern blot hybridization showed that the expression of these genes is induced upon infection with Fusarium graminearum. The accumulation of transcripts for these PR-proteins was more rapid in the resistant variety Sumai 3 than in its susceptible mutant during the first 24 h. This is the first report of the induction of class-IV and class-VII chitinases in cereals by a fungal pathogen.