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Frontiers of Agriculture in China

ISSN 1673-7334

ISSN 1673-744X(Online)

CN 11-5729/S

Front Agric Chin    2009, Vol. 3 Issue (2) : 159-163     DOI: 10.1007/s11703-009-0040-z
RESEARCH ARTICLE |
Biological characteristics of Bacillus thuringiensis strain Bt11 and identification of its cry-type genes
Tinghui LIU1, Wei GUO1,2(), Weiming SUN2, Yongxiang SUN2
1. College of Plant Protection, Agricultural University of Hebei, Biological Control Center of Plant Diseases and Plant Pests of Hebei Province, Baoding 071001, China; 2. College of Life Sciences, Agricultural University of Hebei, Baoding 071001, China
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Abstract  

A novel strain of Bacillus thuringiensis Bt11, isolated from soil samples in China, was classified and characterized in terms of its crystal proteins, cry genes content. The Bt11 strain showed high toxicity against Spodoptera exigua and Helicoverpa armigera neonates. Bt11 strain shares morphological and biochemical characteristics with the previously described Bacillus thuringiensis subsp. kurstaki. SDS-polyacrylamide gel electrophoresis revealed that crystals were composed of several polypeptides ranging from 20 to 130 kDa, of which the 35, 80, and 130 kDa proteins were the major components. PCR-RFLP with total DNA from strain Bt11 and specific primers for cry1, cry2, cry3, cry4/10, cry7, cry8, cry9, and cry11 genes revealed that cry1Aa, cry1Ab, cry1Ia, and cry9Ea genes were present.

Keywords Bacillus thuringiensis      insecticidal activity      biochemical reaction      PCR-RFLP     
Corresponding Authors: GUO Wei,Email:guowei@hebau.edu.cn   
Issue Date: 05 June 2009
URL:  
http://academic.hep.com.cn/fag/EN/10.1007/s11703-009-0040-z     OR     http://academic.hep.com.cn/fag/EN/Y2009/V3/I2/159
colonynumber of larvae testedtreatment concentration/spores·mL-1corrected mortality/%
Helicoverpa armigera505.9×107100
Spodoptera exigua505.9×107100
Tab.1  Bioassay results of Bt11 strain
Fig.1  The spore and crystal of Bt11 strain
strainmyco-demaV.Preactionlecithinasesalicinsurcrosecellobiosemannosestarchhydrolysisproteinhydrolysisurease
Bt11-+++-++++
Tab.2  Physiological and biochemical characteristics of Bt11 strain
Fig.2  The growth curve of Bt11 strain and standard strain HD-73
Fig.3  SDS-PAGE analysis of isolate Bt11
Note: 1 represents protein molecular weight marker (High) (212, 116, 97, 66, 44 kDa); 2 represents Bt11 strain.
geneprimerproduct size/bprestriction enzymeRFLP results/bp
cry1AaK5un2 /K3un21635Pst I & Xba I1117, 518
K5un3 /K3un31463EcoR I& Pst I726, 493, 244
cry1AbK5un2 /K3un21557Pst I & Xba I1039, 518
K5un3 /K3un31463EcoR I & Pst I726, 493, 244
cry1IaS5uni /S3uni1584Bsp119 I & Ban I141, 444, 571, 47, 381
cry2AbS5un2 /S3un21231Hinc II & Msp I791, 297, 143
cry9EaS5un9 /S3un91550Msp I & ClaI1402, 148
Tab.3  -type gene contents of the isolate Bt11
Fig.4  Garose gel eletrophoresis of PCR-RFLP
Note: (a) represents the result of Bt11 and gene PCR-RFLP, with M represents ΦX174- III digest DNA Maker; 1, 2, 3, 4, 5 and 6 representative of Bt11-K2PCR/ I & I, Bt11-K2PCR, Bt11-K3PCR /R I & I, Bt11-K3PCR, Bt11-S2PCR/c II & I and Bt11-S2PCR, respectively. (b) represents the result of Bt11gene PCR-RFLP with M, 1 and 2 representing 100 bp DNA marker, Bt11-S1iPCR/119 I & I and Bt11-S1Ipcr, respectively. (c) represents the result of Bt11 gene PCR-RFLP with M, 1 and 2 representing DNA marker DL2000, Bt11-S9PCR and Bt11-S9PCR/ I & I, respectively.
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