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Frontiers of Agriculture in China

ISSN 1673-7334

ISSN 1673-744X(Online)

CN 11-5729/S

Front Agric Chin    2011, Vol. 5 Issue (3) : 299-304     DOI: 10.1007/s11703-011-1085-3
RESEARCH ARTICLE |
Cloning and pharmaceutical analysis of CaMK gene of Botrytis cinerea
Yuxia DONG1, Jihong XING1, Jiao JIA1, Qiaoyun WENG2, Zhimin HAO1, Jingao DONG1()
1. Molecular Plant Pathology Lab, Agricultural University of Hebei, Baoding 071001, China; 2. Department of Agricultural and Forest Technology, Hebei North University, Zhangjiakou 075131, China
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Abstract  

Using a PCR homology approach, DNA and cDNA sequences of calcium/calmodulin-dependent protein kinase (CaMK) gene of Botrytis cinerea were obtained. Southern blotting result displayed that CaMK was single copy in the genome of B. cinerea. The cDNA sequence of CaMK revealed an open reading frame of 2190 nucleotides encoding a 730 amino acid protein with predicted molecular weight of 81.8748 kDa. The genomic sequence of CaMK revealed the same ORF interrupted by six introns. Bioinformatics analysis showed that this protein had the distinctive features that characterize CaMK ATP binding region signature and serine/threonine protein kinase active-site signature. Pharmaceutical analysis displayed that the CaMK specific inhibitor, KN-62, could inhibit conidial germination, pathogenicity and herbicidal activity of B. cinerea BC4 strain. It was suggested that CaMK played an important role in regulating conidial germination, pathogenicity and herbicidal activity of B. cinerea.

Keywords Botrytis cinerea      CaMK      pharmaceutical analysis      KN-62     
Corresponding Authors: DONG Jingao,Email:shmdjg@hebau.edu.cn   
Issue Date: 05 September 2011
URL:  
http://academic.hep.com.cn/fag/EN/10.1007/s11703-011-1085-3     OR     http://academic.hep.com.cn/fag/EN/Y2011/V5/I3/299
Fig.1  PCR amplification of full-length , cDNA and DNA of BC4. A is PCR amplification of cDNA. B is PCR amplification of DNA.
Fig.2  Conserved domain analysis of CaMK
Fig.3  Predicted secondary structure for CaMK protein from . Blue, red, green and purple represent alpha helix, extended strand, beta turn and random coil, respectively.
Fig.4  Southern Blotting of in the genome of . BC4.
Fig.5  The inhibition of KN-62 on the conidial germination of . . A—C represent concentrations of KN-62 at 60 μmol/L, 80 μmol/L and control, respectively.
Fig.6  The inhibition of KN-62 on the pathogenecity of . . A: . mycelia plugs without KN-62 treatment. B: . mycelia plugs with 80 μmol/L KN-62 treatment. C: Lesion areas were measured 3 d after inoculation. Values are averages from at least eight lesions. Asterisks indicate statistically significant differences between control and treatment, according to Student’s -test.
Fig.7  The herbicidal activity of the toxin of BC4 treated with various KN-62 concentrations. A-E are concentrations of KN-62 at 0 μmol/L, 20 μmol/L, 60 μmol/L, 80 μmol/L and control, respectively.
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