• 식물조직배양학회지
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• 제26권3호
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• pp.177-182
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• 1999

Chitinase 유전자가 삽입된 감자 Belchip 품종의 형질전환체와 대조구에 곰팡이 병원균을 접종하였다. 7개 계통의 형질전환식물체를 12cm정도 키운 후 병원균인 Phytophthora infestans의 zoospore를 접종하여 인공적으로 역병을 유발시켰다. 그 결과 발병율에 따라서 세 그룹으로 분리되었는데 대조구에 비하여 감염 정도가 심한 것 2개 계통. 비슷한 3개 계통, 발병 정도가 약한 2개 계통으로 구분되었다. 대조구에 비하여 저항성이 높았던 2개 계통과 발병이 심했던 1개 계통만을 대상으로 하여 2차실험을 실시한 결과, 1차실험과 비슷한 경향을 나타냈다. 포장에서 생육된 2개의 저항성 계통은 자연적으로 발생한 역병에 대해서도 역시 대조구에 비하여 역병저항성이 더 높았다.

• Plant Biotechnology Reports
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• 제2권1호
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• pp.13-20
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• 2008

Potato (Solanum tuberosum L.), one of the most important food crops, is susceptible to a number of devastating fungal pathogens in addition to bacterial and other pathogens. Producing disease-resistant cultivars has been an effective and useful strategy to combat the attack of pathogens. Potato was transformed with Agrobacterium tumefaciens strain EHA101 harboring chitinase, (ChiC) isolated from Streptomyces griseus strain HUT 6037 and bialaphos resistance (bar) genes in a binary plasmid vector, pEKH1. Polymerase chain reaction (PCR) analysis revealed that the ChiC and bar genes are integrated into the genome of transgenic plants. Different insertion sites of the transgenes (one to six sites for ChiC and three to seven for bar) were indicated by Southern blot analysis of genomic DNA from the transgenic plants. Expression of the ChiC gene at the messenger RNA (mRNA) level was confirmed by Northern blot analysis and that of the bar gene by herbicide resistance assay. The results obviously confirmed that the ChiC and bar genes are successfully integrated and expressed into the genome, resulting in the production of bialaphos-resistant transgenic plants. Disease-resistance assay of the in vitro and greenhouse-grown transgenic plants demonstrated enhanced resistance against the fungal pathogen Alternaria solani (causal agent of early blight).

• BMB Reports
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• 제44권3호
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• pp.193-198
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• 2011

The chitinase-producing strain SC081 was isolated from Korean traditional soy sauce and identified as Bacillus atrophaeus based on a phylogenetic analysis of the 16S rDNA sequence and a phenotypic analysis. A gene encoding chitinase from B. atrophaeus SC081 was cloned in Escherichia coli and was named SCChi-1 (GQ360078). The SCChi-1 nucleotide sequences were composed of 1788 base pairs and 596 amino acids, which were 92.6, 89.6, 89.3, and 78.9% identical to those of Bacillus subtilis (ABG57262), Bacillus pumilus (ABI15082), Bacillus amyloliquefaciens (ABO15008), and Bacillus licheniformis (ACF40833), respectively. A recombinant SCChi-1 containing a hexahistidine tag at the amino-terminus was constructed, overexpressed, and purified in E. coli to characterize SCChi-1. $H_6SCChi$-1 revealed a hydrolytic band on zymograms containing 0.1% glycol chitin and showed the highest lytic activity on colloidal chitin and acidic chitosan. The optimal temperature and pH for chitinolytic activity were $50^{\circ}C$ and pH 8.0, respectively.

• Journal of Microbiology and Biotechnology
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• 제16권6호
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• pp.952-960
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• 2006

As a unicellular green alga that possesses many of the metabolic pathways present in higher plants, Chlorelia offers many advantages for expression of heterologous proteins. Since strong and constitutive promoters are necessary for efficient expression in heterologous expression systems, the development of such promoters for use in the Chlorella system was the aim of this study. Proteins encoded by the early genes of algal viruses are expressed before viral replication, probably by the host transcriptional machinery, and the promoters of these genes might be useful for heterologous expression in Chlorella. In this study, putative promoter regions of DNA polymerase, ATP-dependent DNA ligase, and chitinase genes were amplified from eight Korean Chlorella virus isolates by using primer sets designed based on the sequence of the genome of PBCV-1, the prototype of the Phycodnaviridae. These putative promoter regions were found to contain several cis-acting elements for transcription factors, including the TATA, CAAT, NTBBF1, GATA, and CCAAT boxes. The amplified promoter regions were placed into Chlorella transformation vectors containing a green fluorescence protein (GFP) reporter gene and the Sh ble gene for phleomycin resistance. C. vulgaris protoplasts were transformed and then selected with phleomycin. The GFP fluorescence intensities of cells transformed with chitinase, DNA polymerase, and DNA ligase gene promoter-GFP fusion constructs were 101.5, 100.8, and 95.8%, respectively, of that of CaMV 35S-GFP-transformed Chlorella cells. These results demonstrate that these viral promoters are active in transformed Chlorella.

• 대한약학회:학술대회논문집
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• 대한약학회 2000년도 NEW STRATEGY FOR DRUG DEVELOPMENT IN POST-GENOMIC ERA(대한약학회)
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• pp.205.2-206
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• 2000

• Journal of Microbiology
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• 제45권5호
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• pp.422-430
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• 2007

Clonostachys rosea (syn. Gliocladium roseum) is a well-known biocontrol agent and widely distributed around the world. In this study, an endochitinase gene Crchi1 was isolated from the mycoparasitic fungus C. rosea using the DNA walking strategy. The Crchi1 ORF is 1,746 bp long and interrupted by three introns. The cloned gene Crchi1 encodes 426 amino acid residues and shares a high degree of similarity with other chitinases from entomopathogenic and mycoparasitic fungi. Several putative binding sites for transcriptional regulation of Crchi1 in response to carbon (5'-SYGGRG-3') and nitrogen (5'-GATA-3') were identified in the upstream of Crchi1. Expression of Crchi1 gene in different carbon sources was analyzed using real-time PCR (RT-PCR). We found that the Crchi1 expression was suppressed by glucose but strongly stimulated by chitin or solubilized components of the cell wall from Rhizoctonia solani. Phylogenetic analysis of chitinases from entomopathogenic and mycoparasitic fungi suggests that these chitinases have probably evolved from a common ancestor.

• Journal of Microbiology and Biotechnology
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• 제12권6호
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• pp.865-871
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• 2002

A bacterium having strong chitinolytic activity on $0.2\%$ colloidal chitin-containing agar medium was isolated from coastal soil in Korea. Based on the nucleotide sequence of conserved segment of a 165 rRNA gene, the bacterium was identified as Paenibacillus illinoisensis KJA-424. The population of P. illinoisensis KJA-424 and chitinase activity significantly increased for the first 2 days of incubation. On SDS-PACE analysis with $0.01\%$ glycol chitin, three protein bands (63, 54, and 38 kDa) with chitinolytic activity were detected tooted. The effect of P illinoisensis KJA-424 on the egg hatch of root-knot nematode (Meloidogyne incognita) was investigated. After 7 days of incubation with the chitinase-producing P. illinoisensis KJA-424, none of the eggs hatched, whereas a $39.8\%$ egg hatching rate was observed in the water control. Inverted and scanning electron microscopic observations demonstrated that P. illinoisensis KJA-424 deformed and destroyed the eggshell of M. incognita. In conclusion, chitinase-produced by p. illinoisensis KJA-424 caused the lysis of M. incognita eggshell and resulted in the inhibition of egg hatching in vitro.

• Plant Biotechnology Reports
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• 제3권2호
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• pp.147-155
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• 2009

Oryza grandiglumis Chitinase IVa (OgChitIVa) cDNA encoding a class IV chitinase was cloned from wild rice (Oryza grandiglumis). OgChitIVa cDNA contains an open reading frame of 867 nucleotides encoding 288 amino acid residues with a predicted molecular weight of 30.4 kDa and isoelectric point of 8.48. Deduced amino acid sequences of OgChitIVa include the signal peptide and chitin-binding domain in the N-terminal domain and conserved catalytic domain. OgChitIVa showed significant similarity at the amino acid level with related monocotyledonous rice and maize chitinase, but low similarity with dicotyledoneous chitinase. Southern blot analysis showed that OgChitIVa genes are present as two copies in the wild rice genome. It was shown that RNA expression of OgChitIVa was induced by defense/stress signaling chemicals, such as jasmonic acid, salicylic acid, and ethephon or cantharidin and endothall or wounding, and yeast extract. It was demonstrated that overexpression of OgChitIVa in Arabidopsis resulted in mild resistance against the fungal pathogen, Botrytis cinerea, by lowering disease rate and necrosis size. RT-PCR analysis showed that PR-1 and PR-2 RNA expression was induced in the transgenic lines. Here, we suggest that a novel OgChitIVa gene may play a role in signal transduction process in defense response against B. cinerea in plants.

• 한국응용곤충학회:학술대회논문집
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• 한국응용곤충학회 2002년도 추계 합동 특강 및 학술발표대회
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• pp.138-138
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• 2002

• 한국잠사학회:학술대회논문집
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• 한국잠사학회 2002년도 Join Meetings of Korean Society of Sericultural Science and Japanse Society of Sericultural Science
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• pp.145-145
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• 2002

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