PCR-based Characterization of Bacillus thuringiensis Isolated from Middle Tennessee

Wedad Omar Hawsawi, Tennessee State University


Bacillus thuringiensis(Bt) is a soil-dwelling bacterium that produces an insecticidal crystal protein (delta endotoxin) and vegetative insecticidal protein (Vip) during sporulation and is the most widely used bacterial bio-pesticide due to its toxicity to different insect species. Crystal formation is encoded in plasmid-borne crygenes that can be detected by polymerase chain reaction (PCR). The aim of this study was to detect known and novel cry gene sequences in our stock of 74 Bt isolates collected from Middle Tennessee soil, dust from grain storage and water/soil interfaces by PCR amplification. Two groups of the universal primers were used. The first group had the ability to amplify all the known crygenes and find new ones. The second group had the ability to amplify specific cry gene families such as the known cry1, cry2, cry3, cry4, cry7, cry8 and cry11genes. Forty four (44) of the 47 processed strains were found to contain at least one cry gene when the second group of the primers was used. The most abundant gene was cry1 (87%) followed by cry4(40%), cry 11(38%), and cry2(28%), but none of them were positive for cry3, cry7, and cry 8. Some isolates contained combinations of two or more cry genes; the percentage of the isolates carrying two types of crygenes were12% cry1+cry2,11% cry1+cry4,and 4% cry11+cry4. On the other hand, 11% contained cry1+cry2+cry11 and 17% contained cry1+cry11+cry4, which are three different cry genes. In addition, 4% were positive for four crygenes (cry1+cry2+cry4+cry11), and 30% of the strains possessed a single cry gene. Only three strains were negative with all the primers, which may indicate that these strains may contain different cry genes from the ones that were examined in this study or may contain a uniquecrygene that was not detected by the primers for the known sequences used. The genetic characterization of Cry proteins in this study offers a good opportunity for the selection of the appropriate Bt strains that may have a wider spectrum of activity for agricultural insect pests control as well as control of vector-borne diseases.

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Recommended Citation

Wedad Omar Hawsawi, "PCR-based Characterization of Bacillus thuringiensis Isolated from Middle Tennessee" (2014). ETD Collection for Tennessee State University. Paper AAI1567567.